Active and Adaptive Materials
October 22nd and 23rd 2015, ASRC
1. Velocity Fluctuations in Kinesin-Microtubule Gliding Motility Assays originate from Variations in Motor Attachment Geometry
Henri Palacci*, Ofer Idan, Megan J. Armstrong, Ashutosh Agarwal, Takahiro Nitta, and Henry Hess
Department of Biomedical Engineering, Columbia University
Motor proteins such as kinesin or myosin play a major role in cellular cargo transport, muscle contraction, and cell division. Biomimetic active gels have recently been engineered using kinesin motors and microtubules or myosin motors and actin filaments. Understanding the geometry and collective behavior of coupled motors is critical for our understanding of these systems. An excellent model system is the gliding motility assay, where hundreds of surface-adhered motors propel one cytoskeletal filament such as microtubules or actin. The filament motion can be observed using fluorescence microscopy, revealing small fluctuations in gliding velocity. These observed fluctuations have been previously quantified by a motional diffusion coefficient[3,4]. Sekimoto and Tawada explained the velocity fluctuations as a consequence of the addition and removal of motors from the linear array of motors propelling the filament as it advances, assuming that the motors are not equally effective in propelling the filament. They found that the motional diffusion coefficient is proportional to the motor spacing and the velocity, with a factor of proportionality given by the heterogeneity of motor effectiveness.
Here we combine the measurement of the motional diffusion coefficient with a measurement of the motor spacing and the velocity, which enables us to determine this factor of proportionality in a kinesin/microtubule motility assay. A constant of 0.73 +/- 0.26 (SEM) is measured. We then model the heterogeneity of motor effectiveness arising from the combination of anharmonic tail stiffness and varying attachment geometries (Fig. 1) and obtain good agreement between the factor of proportionality implied by the model and the experimental result. We have thus shown that the work done by a motor varies significantly between motors despite equal energy consumption, leading to an overall loss of effectiveness. This general principle linking effectiveness to the minimization of heterogeneity can be seen in vivo, in the almost perfect alignment of thick and thin filaments in striated muscle.
 F. C. Keber, E. Loiseau, T. Sanchez, S. J. DeCamp, L. Giomi, M. J. Bowick, et al., "Topology and dynamics of active nematic vesicles," Science, vol. 345, pp. 1135-1139, Sep 5 2014.
 J. Alvarado, M. Sheinman, A. Sharma, F. C. MacKintosh, and G. H. Koenderink, "Molecular motors robustly drive active gels to a critically connected state," Nature Physics, vol. 9, pp. 591-597, Sep 2013.
 Y. Imafuku, Y. Y. Toyoshima, and K. Tawada, "Fluctuation in the microtubule sliding movement driven by kinesin in vitro," Biophys J, vol. 70, pp. 878-86., 1996.
 T. Nitta and H. Hess, "Dispersion in Active Transport by Kinesin-Powered Molecular Shuttles," Nano Letters, vol. 5, pp. 1337-1342, 2005.
 K. Sekimoto and K. Tawada, "Extended time correlation of in vitro motility by motor protein," Physical Review Letters, vol. 75, pp. 180-183., 1995.
2. Discovery of Tri-peptide Emulsifiers Using Combined Computational Screening and Experimental Validation
Gary G. Scott, Tell Tuttle and Rein V. Ulijn
Department of Pure and Applied Chemistry, University of Strathclyde
Peptide nanomaterials are an important class of material for the food, cosmetic and biomedical industries. Unfortunately, the self-assembling nature of peptides is often hard to predict. The use of very short (e.g. di- and tri-) peptides has advantages of cost, scalability and rational tenability however have been largely restricted to hydrophobic dipeptides, such as FF. We previously reported the use of coarse-grained molecule dynamics (CG MD)[2-3] as a tool to predict self-assembly behaviour, which led to the discovery of a new class unprotected tripeptide gelators: KYF, KYY, KYW and KFF. The focus of the current work is to further develop this tool coarse-grained molecular dynamics to show that the introduction of organic solvents will allow the creation of emulsified systems. Results show that these tripeptide molecules can act as surfactants, where they assemble at the interface between the octane and water. Experimental methods, such as confocal microscopy, can allow the tracking of these systems, where labelling of the organic solvent with a fluorescent dye allows visualisation of the emulsion system. In addition, spectroscopic analysis (FTIR, fluorescence) is used to assess the peptide arrangements in the emulsions. We have therefore shown that CG MD can also be used for the identification of new emulsifiers comprised wholly of short unprotected peptides.
 Gazit, E et al, Science, 2003, 625-627
 Frederix, P.J.W.M et al, Nat Chem, 2014, 7, 30-37
 Marrink, S et al, J Phys Chem, 2007, 111, 7812-7824
3. Can Supramolecular Gels Be At Thermodynamic Equilibrium?
Ivan Ramos Sasselli*, Tell Tuttle, Peter J. Halling and Rein V. Ulijn
Department of Pure and Applied Chemistry, University of Strathclyde
Low Molecular Weight Gelators are able to form nanostructures, typically fibers, which entangle to form gel-phase materials. These materials have wide-ranging applications in biomedicine and nanotechnology. While it is known that supramolecular gels often represent metastable structures due to the restricted molecular dynamics in the gel state, the thermodynamic nature of the nanofibrous structure is not well understood. Clearly, 3D extended structures will be able to form more interactions than 1D structures. However, self-assembling molecules are typically amphiphilic, thus giving rise to a combination of solvophobic and solvophilic moieties where a level of solvent exposure at the nanostructure surface is favorable. In this study, we introduce a simple packing model, based on prisms with faces of different nature (solvophobic and solvophilic) and variable interaction parameters, to represent amphiphile self-assembly. This model demonstrates that by tuning shape and 'self' or 'solvent' interaction parameters either the 1D fiber or 3D crystal may represent the thermodynamic minimum. The model depends on parameters that relate to fea-tures of experimentally known systems: the number of faces exposed to the solvent or buried in the fiber; the overall shape of the prism; and the free energy penalties associated with the interactions can be adjusted to match their chemical nature. The model is applied to describe the pH dependent gelation/precipitation of well-known gelator Fmoc-FF. We conclude that, despite the fact that most experimentally produced gels probably represent metastable states, one-dimensional fibers can represent thermodynamic equilibrium. This conclusion has critical implications for the theoretical treatment of gels.
4. Localized Self-assembly of Aromatic Peptide Amphiphiles by Immobilized Enzymes
Maria Paola Conte*, K. H. Aaron Lau and Rein V. Ulijn
Department of Pure and Applied Chemistry, University of Strathclyde
Self-assembly of molecular building blocks has attracted increasing interest as an effective bottom-up approach for the design of functional nanomaterials. Amongst the variety of known molecular building blocks, peptides and peptide derivatives attracted particular attention thanks to the huge number of structures that can be achieved through the combination of different amino acid sequences and to the inherent possibility to provide a functional interface with biological materials. Combining (bio-)catalysis and molecular self-assembly provides an effective approach for developing smart biomaterials, allowing to integrate the processes of biological systems with the formation of hierarchical nanostructures. Enzymes proved to be effective to control the self-assembly of aromatic peptide amphiphiles and to develop complex next-generation nanomaterials with possible applications in biology and medicine-related fields. This approach can allow for a highly controlled process in which the assembly is localized in the vicinity of the enzyme. Moreover, with this approach a number of structurally diverse networks can be accessed, depending on the enzyme and precursors selected [2,3]. In this study, the possibility to trigger the localized self-assembly of aromatic peptide amphiphiles exploiting immobilized enzymes is investigated. Catechol chemistry (polydopamine) is identified as a suitable approach to immobilize enzymes on a glass substrate. Protease enzyme thermolysin is immobilized on this substrate. A fluorimetric assay based on Förster resonance energy transfer (FRET) is employed to confirm that the enzyme retains its activity upon immobilization. A soft-lithographic technique (microcontact printing) is used to transfer a pattern of enzymes on a substrate. The surfaces with the patterned thermolysin are employed to trigger the localized formation of the gelator Fmoc-TF-NH2 by reverse hydrolysis of two non-assembling precursors Fmoc-T and F-NH2. Fluorescence microscopy is employed to confirm the localized formation of self-assembled structure by staining the β-sheet fibres with Thioflavin T.
 M. Zelzer and R. V. Ulijn, Chemical Society Reviews, 2010, 39, 3351-3357.
 A. R. Hirst, S. Roy, M. Arora, A. K. Das, N. Hodson, P. Murray, S. Marshall, N. Javid, J. Sefcik, J. Boekhoven, J. H. van Esch, S. Santabarbara, N. T. Hunt and R. V. Ulijn, Nature Chemistry, 2010, 2, 1089-1094.
 R. J. Williams, A. M. Smith, R. Collins, N. Hodson, A. K. Das and R. V. Ulijn, Nature Nanotechnology, 2009, 4, 19-24.
5. Calculating the properties of dynamic covalent peptide macrocycles
Pim W.J.M. Frederix*, Peter C. Kroon, Siewert-Jan Marrink, Sijbren Otto
Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen
Short self-assembling peptides and their derivatives form a useful toolkit to build biocompatible materials. One route towards adaptable materials is Dynamic Combinatorial Covalent Chemistry (DCCC), in which environmental factors or supramolecular interactions direct the equilibrium towards the desired state. Peptide macrocycles, based on disulfide exchange chemistry constitute a particular example of such a system, and demonstrates evolving nanostructures (Science 2010, 327, 1502, Nat. Nanotechnol. 2015, 10, 111). To understand the molecular subtleties that shift the equilibrium we have employed multiscale Molecular Dynamics (MD) and Quantum Mechanical (QM) calculations. These were set up specifically to ease experimental validation (infrared spectroscopy, circular dichroism spectroscopy, HPLC and microscopy) and to aid the design of new biomaterials from libraries of compounds. I will demonstrate a range of relevant computational models and discuss their merits and limitations in the context of peptide macrocycles.
6. Enzymatically activated emulsions stabilized by interfacial self-assembled structures
Inês Moreira*, Tell Tuttle, Rein Ulijn
Department of Pure and Applied Chemistry, University of Strathclyde
In this work, the biocatalytic self-assembly of Fmoc (9-fluorenylmethoxycarbonyl) dipeptide amphiphiles is used in aqueous/organic mixtures to create on-demand emulsifiers. Aromatic peptide amphiphiles have been extensively studied due to their ability to self-assemble into nanostructures through non-covalent interactions, forming self-supporting gels. An alkaline phosphatase is used to transform phosphorylated precursors into self-assembling aromatic peptide amphiphiles, providing a route to trigger self-assembly of nanofibrous networks and hydrogels. The advantages of self-assembled network formation at interfaces are then combined with enzymatic self-assembly to achieve switchable emulsifiers. In particular, the phosphatase-mediated conversion of a phosphorylated dipeptide amphiphile with modest chloroform-in-water emulsion stabilisation capability to the corresponding dephosphorylated gelator (Fmoc-tyrosine-leucine, Fmoc-YL), which forms a stable, permanent interfacial network is described. This gives rise to the possibility of on-demand activation of emulsifying ability, producing switchable emulsions that may be activated by enzyme addition, even after storage of the biphasic mixture for several weeks. Experimental (Fluorescence and FTIR spectroscopy) and computational techniques (Atomistic Molecular Dynamics) are combined to show that the self-assembly process of Fmoc-YL occurs through aromatic interactions and hydrogen bonding to generate a nanostructures network at the water/organic solvent interface.
7. Force controls the material properties of self-assembling branched actin networks: a microscopic analog of 'Wolff's Law'
Tai-De Li*, Peter Bieling, Pamela Jreij, Dyche R. Mullins, and Daniel A. Fletcher
CUNY Advanced Science Research Center
Branched actin networks generate and transmit forces required for a variety of biological processes but we do not understand what controls the mechanical properties of these self-assembling materials. Using atomic force microscopy we measured the material properties of branched actin networks assembled in vitro under physical boundary conditions that mimic those experienced in cells. Surprisingly, we discovered that the material properties of branched actin networks are not well described by existing physical theories. In addition, we found that networks grown under high loads are stronger and stiffer than those assembled under low loads, and that branched actin networks are stiffest and exhibit the least internal friction when loaded at their growth force. Exposing assembled networks to higher or lower loads either irreversibly damages them or causes them to soften elastically. Branched actin networks, therefore, exhibit memory and dynamically adapt their material properties to best resist the load they experience during self-assembly. Adaptation and memory also uniquely determine the force-velocity relationships of branched actin networks assembled under changing stresses. Our results reveal a microscopic, cytoskeletal analog of Wolff's law, which describes how force enhances the density and stiffness of trabecular bone in the vertebrate skeleton
8. Transport-Mediated Molecular Weight Distributions in Self-Assembling Amphipathic Peptides Grown Near Interfaces
Matthew B. Kubilius* and Raymond S. Tu
Department of Chemical Engineering, City College of New York
Synthesizing a periodically-sequenced, useful, amphipathic peptide is challenging due to the polydispersity index increases of large molecular weight polypeptide systems. To overcome this, we designed synthetic amino acid dimers that are both amphipathic and water-soluble. When polymerized, these dimers give rise to a peptide with alternating hydrophilic/hydrophobic side groups: the typical periodicity for beta-sheet forming polypeptides. Using this approach, we can influence polydispersity in the growing polypeptide chains, controlling the kinetics of growth through transport-limited chain elongation. Our experiments show that in the absence of a micellular interface, standard bulk-phase condensation polymerization occurs. The amphipathic character of the peptide chain increases with increasing molecular weight, resulting in a polypeptide that partitions into surfactant micelles as a function of the degree of polymerization. This type of kinetically-limited growth serves to narrow the polydispersity of our periodically-sequenced polypeptide. We quantify the dynamics of chain elongation and interfacial assembly using multi-angle light scattering and mass spectrometry and define the evolving sheet-like secondary structure using circular dichroism for various peptides of differing amino acid pairs. Our results show that the peptides grown in the presence of micelles show significantly enhanced self-assembly and a narrowed polydispersity index. From this, we conclude that the transport-limited chain elongation polymerization method shows great promise in the manufacture of low-cost, interfacially assembling polypeptides.
9. Molecular Modeling of Supramolecular Assembly of Drug Amphiphiles
Myungshim Kang,* Pengcheng Zhang, Honggang Cui, Sharon M. Loverde
Department of Chemistry, College of Staten Island, CUNY
Recently, drug amphiphiles (DAs) have been shown to form discrete and stable supramolecular nanostructures with high and quantitative drug loading1. A drug amphiphile consists of a hydrogen-bonding peptide sequence attached to a hydrophobic drug. Similar to peptide amphiphiles2, DAs also self-assemble into discrete and well-defined supramolecular structures. Using molecular dynamics simulations we investigate inter- and intra-molecular interactions driving the self-assembly and formation of the morphology of supramolecular DAs. More specifically, we investigate the self-assembly of camptothecin-based DAs, which have been shown to form cylindrical supramolecular assemblies with a well-defined structure. We examine the self-assembly process from random using long-time all-atomistic molecular dynamics simulations (> 200 ns). We also examine the structure of pre-assembled cylindrical supramolecular assembly, analyzing radial distribution of each component of DAs and Cl- ions in the cylinder. We also characterize π-π stacking using the distribution of distances and angles between the planar CPTs in DA, as well as hydrogen bond formation in the peptide over time. Our findings can help add further insight into the rational design of supramolecular assemblies.
10. Light-emitting self-assembled peptide nucleic acids exhibit both stacking interactions and Watson-Crick base pairing
Or Berger*, Lihi Adler-Abramovich, Michal Levy-Sakin, Assaf Grunwald, Yael Liebes-Peer, Ludmila Buzhansky, Tal Schwartz, Yuval Ebenstein, Felix Frolow, Linda J. W. Shimon & Ehud Gazit
Department of Molecular Microbiology & Biotechnology, Tel Aviv University
The two main branches of bionanotechnology involve the self-assembly of either peptides or DNA. Peptide scaffolds offer chemical versatility, architectural flexibility and structural complexity, but they lack the precise base pairing and molecular recognition available with nucleic acid assemblies. Here, inspired by the ability of aromatic dipeptides to form ordered nanostructures with unique physical properties, we explore the assembly of peptide nucleic acids (PNAs), which are short DNA mimics that have an amide backbone. All 16 combinations of the very short di-PNA building blocks were synthesized and assayed for their ability to self-associate. Only three guanine-containing di-PNAs-CG, GC and GG-could form ordered assemblies, as observed by electron microscopy, and these di-PNAs efficiently assembled into discrete architectures within a few minutes. The X-ray crystal structure of the GC di-PNA showed the occurrence of both stacking interactions and Watson-Crick base pairing. The assemblies were also found to exhibit optical properties including wide-range excitation-dependent fluorescence in the visible region.
11. Normal Mode Analysis of α-β Tubulin Dimers
Anjela Manandhar, Myungshim Kang, Sharon M. Loverde
Department of Chemistry, College of Staten Island
The basic building block of an MT is heterodimer tubulin consisting of α and β subunits. These dimers align longitudinally in head to tail fashion to form polar protofilaments (PFs) and PFs combine laterally to form cylindrical, hollow MT structure. These MTs are 25 nm wide and nm to μm long. Microtubule growth is governed by dynamic instability stochastic switching between polymerization and de-polymerization phases. At the molecular level the dynamics of the MT is driven by the hydrolysis state of GTP in the β subunit of tubulin. In this study, we use normal mode analysis (NMA) to study functionally important global motions of the tubulin dimer. We utilize a model of the α-β tubulin dimer from zinc induced tubulin sheets. NMA results show four distinct modes - twisting, stretching and two bending modes. This compares well with previous studies by Bahar et al. Next, we will conduct NMA for recent high-resolution MT structures of dynamic microtubules by Nogales et al. to compare differences in the global motions with previous studies. These comparisons will provide insight towards structural and dynamical differences between different models of tubulin, as well differences between GTP and GDP states.
12. Molecular Dynamics of Single Chain Hydrophobic Polymers in Water
Department of Physics/Chemistry, CUNY-CSI
The hydrophobic nature of polymers can be characterized by the scaling behavior of single chains in various solvents. In the study, we perform atomistic (AA) molecular dynamics simulations of poly(styrene) (PS) chains in water. We calculate chain properties such as the radius of gyration and end to end distance as a function of monomer length. Following, we perform metadynamics calculations of single polymer chains of PS using radius of gyration as a reaction coordinate.
Next we will compare the results of these simulations with a coarse grained (CG) model of PS.
13. Active Photonic Hypercrystals
T. Galfsky *, E.E. Narimanov, and V. M. Menon
Department of Physics, City College of New York
Photonic crystals and metamaterials are two of the major paradigms in the field of photonics that have resulted in a plethora of discoveries both of fundamental and technological importance. A photonic hyper crystal (PHC) is a metamaterial/photonic crystal hybrid which opens a new universality class of artificial optical media for nanophotonics applications.
We facbricated an active two-dimensional photonic hypercrystal that shows enhanced spontaneous emission from its metamaterial component and light extraction through its photonic crystal property. Spontaneous decay rate of embedded quantum dots is enhanced by a factor of 19.5 and light extraction from the HMM is enhanced by a factor of ~100.
14. Spatiotemporal Profiling the Activities of Ectophosphatases on Live Cells
Jie Zhou*, Xuewen Du, Cristina Berciu, Junfeng Shi, Daniela Nicastro, and Bing Xu†
Department of Chemistry, Brandeis University
(† Corresponding author)
Ectoenzymes, which have the catalytic domains outside the plasma membrane of cells, usually carry out dual or multiple functions in many types of cells. Despite overexpression of ectophosphatases represents a generic difference between certain cancer and normal cells, the evaluation of their activities is rather difficult and receives little attention. Here we report the profiling of the activities of ectophosphatases on cancer cells by enzyme-catalyzed self-assembly of a D-peptidic hydrogelators that form fluorescent molecular nanofibrils. Possessing an environment-sensitive fluorophore and an enzyme substrate (i.e., phosphorylated tyrosine), the designed precursor can turn into the corresponding hydrogelator via enzyme-instructed dephosphorylation, which results in the self-assembly of the hydrogelator and gives enhanced fluorescence. Our study reveals the significantly higher activities of ectophosphatases on cancer cells than on stromal cells in co-culture and shows an inherent and dynamic difference in phosphatase activities between drug sensitive and resistant cancer cells or between cancer cells with and without hormonal stimulation. Besides providing an approach to achieve high spatiotemporal resolution for profiling the activities of phosphatases, a ubiquitous class of enzymes, in mixed population of live cells, the integration of enzyme catalysis with self-assembly serves as a novel approach to profile the activities of enzymes and to control the fate of cells.
15. Demonstration of energy transfer within self-assembling bioelectronic hydrogelators with tunable properties
Herdeline Ann M. Ardoña*, Kalpana Besar, Matteo Togninalli, Brian P. Ginn, Hai-Quan Mao, Howard E. Katz, John D. Tovar
Department of Chemistry, Johns Hopkins University
The engineering of supramolecular assemblies at the molecular level to achieve a specific function has been a longstanding challenge in various fields that utilizes bottom-up self-assembly strategies. We present a coassembled pi-conjugated peptide system composed of oligo-(p-phenylenevinylene)-based donor units and quaterthiophene-based acceptor units that demonstrate energy transfer in completely aqueous environments. The formation of one-dimensional nanostructures by the peptide assemblies bearing semiconducting units encourages energy migration along the stacking axis, thus resulting in the quenching of donor emission peaks along with the development of new spectral features reminiscent of acceptor emission. The effect of environmental stimuli (pH, temperature) to nanostructure formation and energy transfer will also be presented. To further emphasize the ability of these peptide-pi-peptide constructs to have rationally designed properties, the study on the sequence-dependent rheological, photophysical and electrical properties of these semiconducting peptide hydrogelators will be presented. This aspect demonstrates the effects of amino acid sequence on the nanoscale to the macroscale electrical transport and mechanical properties of nanostructure-forming pi-conjugated peptides. With the optimized conditions for the construction of peptide hydrogels with the desired properties, our group is currently utilizing these pi-conjugated peptide hydrogels as scaffolds for human neural stem cells. Overall, this unique material design that can coassemble two different semiconducting units within peptide nanostructures and have sequence-tunable properties offers a new platform for the engineering of energy transport through bioelectronic materials in aqueous environments.
16. Optimal Learning for Novel Materials Discovery
Jialei Wang*, Yingjie Fei, Matthias Poloczek, Lorillee Tallorin, Nicolas Kosa, Matthew Thomspon, Swagat Sahu, Pu Yang, Peter Frazier, Michael Gilson, Nathan Gianneschi, Michael Burkart
Operations Research & Information Engineering, Cornell University
Scientists use laboratory experiments to find and verify materials with desirable properties, e.g., that make efficient solar cells; or that deliver a drug to a target within the human body. The success of these searches hinges on making good decisions about which experiments to perform. We show that machine learning and optimization, together in an optimal learning framework, can be used to choose good experiments, and to reach experimental goals reliably with fewer experiments than alternate techniques. We describe how these techniques were used successfully to find minimal peptide substrates for a pair of protein-modifying enzymes, with application to novel biochemical sensors.
17. Spontaneous emergence of autocatalytic information-coding polymers
Alexei Tkachenko*, Sergei Maslov
Center For Functional Nanomaterials, Brookhaven National Laboratory
Self-replicating systems based on information-coding polymers are of crucial importance in biology. They also recently emerged as a paradigm in design on nano- and micro-scales. I will discuss a general theoretical analysis of the problem of spontaneous emergence of autocatalysis for heteropolymers capable of template-assisted ligation driven by cyclic changes in the environment. Our central result is the existence of the first order transition between the regime dominated by free monomers and that with a self-sustaining population of sufficiently long oligomers. We provide a simple mathematically tractable model that predicts the parameters for the onset of autocatalysis and the distribution of chain lengths, in terms of monomer concentration, and two fundamental rate constants. Template-assisted ligation allows for heritable transmission of information encoded in oligomer sequences thus opening up the possibility of long-term memory and Darwinian evolution of such systems.
This research used resources of the Center for Functional Nanomaterials, which is a U.S. DOE Office of Science Facility, at Brookhaven National Laboratory under Contract No. DE-SC0012704. Work at Biosciences Department was supported by US Department of Energy Office of Biological Research Grant PM-031.
18. Traceable engineered protein block copolymers for hyperthermic drug delivery
Joseph A. Frezzo*, Jin Kim Montclare, Ph.D.
Department of Chemical and Biomolecular Engineering, NYU School of Engineering
Theranostics represent a hybrid field where both drug delivery and medical imaging are merged to promote more effective therapies. In this work, a protein block copolymer, recombinantly expressed in a leucine auxotrophic strain of E. coli for 5,5,5,-l-trifluoroleucine (TFL) incorporation, is investigated for potential theranostic use as a chemotherapeutic carrier and visualized via MRI. The fluorinated protein block copolymer, dubbed CE2-RGD-TFL, is comprised of two functional domains: 1) a coiled-coil domain (C), flanked by two integrin targeting domains, that encapsulates small hydrophobic molecules and; 2) two sequential elastin-like peptide domains (E) that impart concentration dependent thermoresponsiveness2. Incorporation of TFL in CE2-RGD yields a drug carrier nanoparticle that can undergo temperature dependent structural changes measured by an increase in R2 values which lends merit to CE2-RGD-TFL as a potential T2-weighted MRI contrast agent. Also, fluorination imparts greater thermoresponsiveness in the physiological range for hyperthermic drug delivery. Furthermore, the fluorinated protein demonstrates greater loading of doxorubicin which provides further value to CE2-RGD-TFL as a theranostic agent.
19. Synthesis of conductive protein nanofibers and biofilms
Noémie-Manuelle Dorval Courchesne*, Pei Kun R. Tay, Peter Q. Nguyen, Neel S. Joshi
Wyss Institute for Biologically Inspired Engineering, Harvard University
Protein-based materials capable of electron transfer are attractive for several applications, including microbial fuel cells, biosensors, electrobiosynthetic systems, and flexible electronics. While some bacterial species like Shewanella and Geobacter naturally produce conductive nanowires, these nanowires are difficult to engineer via rational design. Here, we propose to use Escherichia coli as a well-known and easily engineerable microorganism to serve as a factory for the production of conductive protein nanofibers. E. coli naturally secretes curli fibers that promote biofilm formation. These fibers are composed of CsgA proteins that self-assemble to form micron-long extracellular amyloid-folded appendages. Using a biomimetic approach, we designed strategies to allow for electron transfer along these curli fibers based on electron delocalization through pi-stacks formed by aromatic amino acids. We genetically engineered E. coli and expressed several mutant CsgA proteins containing series of aligned aromatic amino acids. The mutants can still self-assemble into amyloid fibers as confirmed by Congo red binding assays and electron microscopy. We are investigating the conductive properties of our mutant fibers using electrochemical methods for purified his-tagged fibers, or for bulk living biofilms. Additional functionalities could be introduced in the curli network by fusing CsgA with peptides that allow for binding, enzymatic reactions, or covalent chemical modifications. Overall, our system provides a tunable platform for the synthesis of conductive protein networks and biofilms.
20. Tuning Interparticle Interactions to Control Self-Assembly of DNA-Coated Colloids
Hasan Zerze*, Yajun Ding, Minseok Song, Jeetain Mittal
Department of Chemical and Biomolecular Engineering, Lehigh University
DNA-programmed colloids have gained remarkable interest as building blocks of tunable super-lattice architectures. Hierarchical arrangements of noble metal nanoparticles with the DNA-directed self-assembly promise unique applications in a wide variety of fields such as nano-lasing devices, molecular sensors, and optoelectronics. As a design strategy, the particles can be functionalized with complementary single stranded DNA molecules as either single-flavored (one type of DNA on each particle) or multi-flavored (multiple types of DNA strands on each particle). The multi-flavoring approach can be successfully utilized as a flexible handle for tuning inter-particle interactions.
In the present work, we first use a coarse-grained model of DNA-functionalized particles (DFPs) to calculate particle-particle interactions. The resulting information is used to construct an even simplified model of multi-flavored DFPs to study the spontaneous self-assembly of particles into different crystalline structures. In a two-dimensional system, we find that DNA-coated particles can spontaneously organize into well-defined square and hexagonal lattices as a function of interparticle attraction strength. Additional control over compositional order of particles in honeycomb and kagome structures can also be obtained in this case. We use a similar strategy for three-dimensional assembly of particles and can access a wide range of interesting crystal structures with high degree of compositional order. Our study clearly demonstrates new structural capabilities of DNA-functionalized materials based on their ease of control of pairwise interactions through different design strategies.
21. Supramolecular Polymers and Hydrogels for Biomedical Applications
Lye Lin Lock*, Xinpei Mao and Honggang Cui
Department of Chemical and Biomolecular Engineering, Johns Hopkins University
Supramolecular polymers are polymeric arrays of small molecular building units linked by non-covalent interactions. This non-covalent and living nature provides unique properties that expand the functional space of classical covalent polymers. Our research efforts center on the design and synthesis of small molecular building units to create functional supramolecular polymers and hydrogels for use in drug delivery systems and wound care. The supramolecular polymers of our particular interest are shape persistent one-dimensional (1D) nanostructures with a high degree of internal order. The unique aspect of our strategy is to harness the hydrophobic property of drugs (antibiotics, anti-inflammatory agents, or anticancer drugs) to drive the formation of drug-based supramolecular polymers through the creation of drug amphiphiles (DAs). These DAs can be synthesized by the conjugation of a hydrophilic peptide moiety that gives the resulting molecules overall amphiphilicity. The assembled DAs possess a precisely controlled and high drug loading and will shield the drug from its environment, protecting it from unwanted degradation. Under appropriate conditions these filamentous nanostructures can then enmesh to give a hydrogel, which can be used for regenerative medicine, wound dressings, and local drug delivery.
22. NMR studies of natural and designed elastins
R.L. Koder, Kelly Greenland, Tatiana Krivhokhizhina and Richard Wittebort
Department of Chemistry, University of Louisville
We are investigating protein and solvent dynamics in nature's predominant elastomer, elastin, using a combination of NMR and protein design approaches. Studies of protein dynamics by NMR with site-specific resolution are hampered by elastin's high molecular weight, its sequence complexity and it's random coil structure. To mitigate these problems, we have followed Keeley's 'minielastin' approach and expressed constructs containing tandem repeats of hydrophobic, 'H', and cross-link, 'X', modules. To simplify NMR spectra, 'H' modules contain exact repeats like APGVGV and the 'X' modules contain AKAAKA motifs. NMR spectra of these constructs are sufficiently well resolved for site-specific ssNMR experiments and complete resonance assignments were obtained using standard 3D NMR methods. Early observations include (i) resonance shifts in 'H' modules have random coil values (ii) 'H' module shifts are independent of the flanking module and equivalent to those of a single 'H' module and (iii) 'X' module shifts differ from random coil values and are affected by the presence of flanking 'H' modules. To study the role of hydration in elastic recoil, we use 2Q 2H NMR of stretched fibers equilibrated with 2H2O. This allows us to use multiple quantum filtration to distinguish surface water and bulk water in fully hydrated fibrils. Long ago, Weis-Fogh and Gosline suggested that a significant contribution of hydrophobic forces, i.e., stretch induced ordering of water at the solvent:protein interface, is key to the driving force for recoil. We find an order of magnitude increase in the amount of ordered water when fibrils are stretched 1.5-fold from their relaxed length.
23. Enzyme-responsive minimalistic peptide design: Applications in nanoscience and biology
Jugal Kishore Sahoo1, Duncan Graham2, Mathew Dalby3, Rein V Ulijn1,4
1 Technology and Innovation Centre, University of Strathclyde, Glasgow,
2 Centre of Molecular Nanometrology, WestCHEM, Department of Pure and Applied Chemistry, Technology and Innovation Centre, University of Strathclyde, Glasgow
3 Centre for Cell Engineering, University of Glasgow, Glasgow,
4 Advanced Science Research Centre (ASRC) and Hunter College, City University of New York, NY
Enzyme responsive peptidic materials are a new class of stimuli responsive smart (bio-) materials that undergo supramolecular transition when triggered by the catalytic action of the enzymes. The use of enzymes as an external trigger, and short peptides as versatile building blocks opens up new avenues for a vast range of applications in the field of nanoscience, biology and regenerative medicine.
In here, three different classes of enzyme responsive peptide systems will be demonstrated, 1) Adaptive soft matter through supramolecular network; an azobenzene appended dipeptide unit (Azo-YF-NH2) undergoes gelation and amide condensation in presence of thermolysin. However, when exposed to light of specific wavelength, it undergoes supramolecular reconfiguration because of the photo-isomerisation properties of azobenzene, i.e. trans-azobenezene based dipeptide forms hydrogels and fiber-like morphology whereas cis-azobenzene forms solutions and micellar structures. 2) peptide hydrogels as reductive template for nanoparticle synthesis; aromatic short dipeptide hydrogels (Fmoc-FY) can be used as a reductive template for in situ synthesis and stabilisation of gold nanoparticles. The size of the nanoparticles synthesized depends on the concentration of enzyme used, i.e. higher the concentration of enzyme, smaller is the particle size and vice versa. 3) Dynamic switchable 2D peptide surface; an enzyme responsive 2D peptide surface which changes its chemical properties when exposed to enzyme has been designed, which contains a cell adhesive tripeptide (RGD), an elastase responsive dipeptide unit (AA). Both the peptides are camouflaged by a bulky group (Fmoc- or PEG-). This peptide surface offers user induced control of surface adhesivity of stem cells which in turn allows sequential control of growth and differentiation on demand on a surface, using enzymatic action as a trigger.
 J. K. Sahoo et al., Biocatalytic amide condensation and gelation controlled by light, Chem Commun., 2014, 50, 5462.
 J. K. Sahoo et al., Non-equillibrium biocatalytic self-assembly for templating of gold nanoparticles, under preparation.
 J. K. Sahoo et al., Analysis of enzyme-responsive peptide functionalised surfaces by Raman spectroscopy, under preparation.
 J. N. Roberts, L. E. Mcnamara, J. K. Sahoo et al., Dynamic regulation of mesenchymal stem cell adhesion footprint to control growth and differentiation on-demand, under revision.
24. Engineering the Bacterial Extracellular Matrix for Controlling the Localized Delivery of Anti-inflammatory Factors in the Gastrointestinal Tract
Anna Duraj-Thatte1,2, Pichet Praveschotinunt1,2, Trevor Nash1,2, Fred Ward2,3 & Neel Joshi1,2
1School of Engineering and Applied Sciences, Harvard University, 2Wyss Institute for Biologically Inspired Engineering, Harvard University 3Department of Chemistry, Harvard University
The number of cases of an inflammatory bowel diseases (IBD) such as Crohn's disease and ulcerative colitis as well as gut inflammation have significantly increased in last few decades. Existing therapeutic strategies for IBD treatment are not always effective in maintaining IBD remission. It brings a need for search the new therapies that are not only able to improve quality of patient's life but also help stopping the inflammatory process.
Recent discoveries showing the importance of gut microbiota in human diseases has generated great interest in engineering these microbes. Prior approaches to engineering beneficial microbes have focused mostly on demonstrating their ability to secrete optimum amounts of recombinant protein-based therapeutics; but so far there has been little effort to rationally control the localization of the engineered microbes to specific sites within the gut.
Here we develop a commensal biofilm matrix as a material capable of targeted and triggered biologic delivery to treat inflammatory diseases of the gastrointestinal tract.
Various components of the bacterial extracellular matrix have been implicated in adhesion to the surface of the intestinal epithelium. One of them, the curli fibers, are a major structural component of the E.coli biofilm. These are the amyloid fibers expressed on the surface of E. coli. They are produced through a biosynthetic pathway controlled by the curli operon, and assembled via the extracellular nucleation-precipitation pathway, which allows for secretion the amyloidogenic monomers CsgA into the extracellular space. The CsgA proteins can be altered to create a programmable, biofilm by attaching functional peptides onto the C-terminus of the protein. We hypothesized that engineering the curli matrix by attaching specific peptides fused to CsgA protein could lead to the development of bacteria with enhanced colonization and ability to modulate localized adhesion in the gut. We have engineered the curli of commensal E.coli with displayed trefoil factors (TFFs), a family of small peptides secreted from the GI tract, that play a important role in the protection and the repair of the gastrointestinal epithelial surface. We have shown the ability of the bacteria to express the modified curli fibers with displayed TFFs. Such engineered bacterial extracellular matrix is not only able to form a biofilm with displayed TFFs but also adhere to a human epithelial colorectal carcinoma cell line better than bacteria without displayed peptides. We have also shown that curli bind TFFs are able to enhance cell migration and up or down regulate various biomarkers correlated with TFF activity.
We believe that such a functionalized biofilm not only has a significant potential to enhance the localized colonization of the commensal microflora but also be a programmable platform with immobilizing therapeutic peptides to target the specific inflammation sites in the gastrointestinal tract.
25. Transient pulses and stable gradients in hydrogel microstructures through quasi-permanent actuation schemes
Peter A. Korevaar*, Reanne M. Rust, Joanna Aizenberg
John A. Paulson School of Engineering and Applied Sciences, Harvard University
The reversible swelling and contraction of hydrogels is widely applied to drive the mechanical actuation of functional microstructures. Typically, hydrogel micro-actuators are designed to operate in concert to the actual environmental stimulus (e.g. temperature, humidity, pH, electric field). Here we explore alternative, quasi-permanent actuation schemes based on polyacrylic acid hydrogels. Quasi-permanent actuation allows locally addressing the micro-actuators and thereby creating gradients that remain stable after removal of the trigger - behavior that is relevant to many applications, but impossible to obtain with hydrogel systems that continuously re-equilibrate to their environment. Besides stepwise responses to pulse-like triggers, our system also allows generating transient pulses in the hydrogel swelling/contraction as a response to stepwise triggers. Our approach provides potential application to energy-efficient operation of micro-actuators, and opens avenues for complex actuation routines at the microscale.
Catalytic biofilms by self-assembling enzyme immobilization onto E. coli curli nanofibers
Martin G. Nussbaumer*, Zsofia Botynszki, Pei Kun R. Tay, Peter Q. Nguyen, Neel S. Joshi
Wyss Institute for Biologically Inspired Engineering, Harvard University
Performing stereoselective biocatalytic transformation is of great interest to the pharmaceutical industry. Nowadays biocatalytic transformation relies on whole cells or purified enzymes. However, these systems have inherent disadvantages, such as high costs of enzyme purification or low mass transport in whole cell systems. Our strategy to overcome these problems is based on rationally designed biocatalytic surfaces, the so-called Biofilm Integrated Nanofiber Display (BIND) system. The BIND exploits the protein-nanofiber network of E. coli biofilms, which we modified in such way that enzymes are site-specifically immobilized on the nanofibers. The immobilization of enzymes onto BIND rest upon different protein-peptide interaction with no need for any chemical. This approach was used to immobilize dehydrogenase (DH), which serves in future work as co-factor regeneration for stereoselective ketoreductases in the synthesis of active pharmaceutical ingredient precursors.
27. Sequence Adaptive Peptide-Polysaccharide Nanostructures by Biocatalytic Self-Assembly
Yousef M. Abul-Haija and Rein V. Ulijn
WestCHEM/Department of Pure & Applied Chemistry and Technology & Innovation Centre, University of Strathclyde
Molecular self-assembly coupled with (bio)catalysis underlie dynamic processes in biology and are considered as powerful tools for fabricating adaptive, functional (bio)materials. One approach to achieve dynamic functionality is by using co-assembly (with biomacromolecules). Herein, we aim to develop synthetic mimics of biological systems by the combination of co-assembly and biocatalysis. Peptides are particularly useful as building blocks in the biomaterials context, due to the rich chemistry and functionality available from primary sequences of twenty amino acids, as well as their biological relevance. We show that in situ enzymatic exchange of a dipeptide sequences in aromatic peptide amphiphiles/polysaccharide co-assemblies enables dynamic formation and degradation of different nanostructures depending on the nature of the polysaccharide present. This is achieved in a one-pot system composed of Fmoc-cystic acid (CA), Fmoc-lysine (K) plus phenylalanine amide (F) in the presence of thermolysin which, through dynamic hydrolysis and amide formation gives rise to dynamic peptide library composed of the corresponding Fmoc-dipeptides (CAF and KF). When the cationic polysaccharide chitosan is added to this mixture, selective amplification of the CAF peptide is observed giving rise to formation of nanosheets through co-assembly. By contrast, upon addition of anionic heparin, KF is formed which gives rise to a nanotube morphology. The dynamic adaptive potential was demonstrated by sequential morphology changes depending on the sequence of polysaccharide addition. This first demonstration of the ability to access different peptide sequences and nanostructures depending on presence of biopolymers may pave the way to biomaterials that can adapt their structure and function and may be of relevance in design of materials able to undergo dynamic morphogenesis.
28. DNA Origami Virus-Binding 'Claws': Binding and Antibody Attachment Studies; Making DNA 'Photoswitch' Surfaces Less Prone to 'Leakage'
Ekaterina Selivanovitch*, Christopher Chen, Sydney Snaider, and Philip S Lukeman
Department of Chemistry, St. John's University
We use DNA origami (DO) in order to assemble a macromolecular 'claw' containing regions of single strands complementary to those on a modified virus capsid. Upon binding, claw should undergo a large conformational change. DO 'Claw' is modified to contain various number of binding sites and limited mobility in order to increase the binding efficiency to the modified capsid. We expect these claws to act as prototypes of the key components for a self-contained, low background, portable, electrochemical virus biosensor.
Additionally, DNA-based switches are currently operated by manual, solution-phase addition of 'set-strands'. Here we have sterically inaccessible 'set-strands', released from a surface into solution by spatially controlled photocleavage, that operates both stoichiometric and nucleated set-strand controlled systems. This technique will enable microarrays of set-strands to operate many DNA-based switches in computational and diagnostic devices.
29. Sustainable Molecular Organogels: Green Soft Materials for Functional Next Generation Fuels
Julian R. Silverman*, Dr. George John
Department of Chemistry, CCNY and the CUNY Graduate Center Doctoral Program
A multifunctional small molecule capable of thickening and gelling a variety of liquids has been synthesized from natural renewable resources. The gelator is derived from biodiesel, a waste product and fuel, and successfully gelates mixtures of diesel and biodiesel, which are currently sold as fuel alternatives. The gels of these mixtures prove to be robust (1k-100k Pa) even at exceedingly low (<1% mass/volume, MGC: 0.46 wt.%, Tgel 80 oC) concentrations. These mixtures may serve as potential alternatives to liquid fuels, which are known to spill and cause ecological devastation in environmental disasters. These adaptive materials respond both to temperature, and shear stimuli, and may transition from solid-like gel to liquid solution reversibly. The thixotropic nature of these gels makes them not merely shear responsive, but fully susceptible to shear-induced gelation, an interesting alternative to heating and cooling to form gels. Rheology may be used to probe both the mechanical nature of these systems indicating the presence of a network of fibers, which is confirmed by electron microscopy (100 nm fibers). Using the versatile characteristics of these novel gelator systems they may thus be tuned to afford materials specific to any variety of applications.
30. Mechanical sensing with mechanochromic soft materials
F. Cellini*, S. Khapli, S.D. Peterson, and M. Porfiri
Department of Mechanical and Aerospace Engineering, New York University
Several classes of materials, such as glasses and organic crystals, present optical properties that can be modulated in response to mechanical deformation, opening avenues for the development of new sensing devices and smart systems. Mechanochromic polymers use a fluorescent organic dye molecule to afford a substantial change in coloration in the visible spectrum during mechanical deformation. In this study, we investigate the reversible optomechanical behavior of mechanochromic elastomers. We present results for the experimental characterization of these soft materials during uniaxial and biaxial testing by simultaneous acquisition of the applied load, mechanical deformation, and fluorescence emission. Experimental data are analyzed through rigorous constitutive modelling of both mechanical and optical responses. We expect that this knowledge will aid in the design of strain and stress sensors with applications in mechanics, biomechanics, and life sciences.
31. MMP-9 triggered formation of doxorubicin depots halts tumor growth
Daniela Kalafatovic*, Max Nobis, Kurt I. Anderson and Rein V. Ulijn
Nanoscience Initiative, CUNY ASRC
Expression levels of enzymes dictate the difference between health and disease in many cases, including cancer. This leads to design peptide amphiphiles that upon cleavage by a disease-associated enzyme reconfigure from micellar aggregates to fibres. The designed PhAc-FFAGLDD (1a) and GFFLGLDD (2a) and their expected products of enzyme cleavage PhAc-FFAG (1b) and GFFLG (2b) were synthesized and characterized by AFM, FTIR, DLS, rheology and fluorescence. After the designed peptides were shown to be successful in controlling the morphology of the supramolecular aggregates based on the peptide length i.e. hydrophobicity, the enzyme triggered micelle to fibre transition was explored. Following this it was investigated whether the micelles were capable to perform as mobile vehicles for encapsulation and release of hydrophobic drugs. It was observed that the assembled fibres provide a new scaffold for prolonged drug delivery due to the partial entrapment of the drug and the intrinsic biodegradable nature of peptide carriers themselves. Being purely peptidic these systems have the advantage of being not toxic to cells (MTT assay) and can be used as carriers for doxorubicin in vivo. When tested on animal models, the cancer growth in slowed down by administration of doxorubicin loaded peptides compared to doxorubicin only. This approach opens up the possibility of developing new (enzyme responsive) drug delivery vehicles designed to release drug payload locally, in the presence of the desired enzyme with potential application in cancer therapy.
32. Reactions in Individual Droplets on a Superhydrophobic Surface: Effect of Convection
Yang Liu*, Xiaoxiao Chen, Qian Feng Xu, Alexander Greer, Yuanyuan Zhao, and Alan M. Lyons
Department of Chemistry, College of Staten Island, CUNY
On a superhydrophobic surface, an aqueous droplet can maintain its quasi-spherical shape without wetting the surface. This geometry creates a nearly isothermal micro-scale container in which chemical reactions and analysis can be conducted using small volumes (<10 μL) of reagents. However, due to their small size and thermal isolation from the substrate, convection in these droplets is suppressed and so mixing within these micro-containers occurs only by diffusion. In this paper, we present a method to induce and control convective mixing within small size droplets poised on multifunctional superhydrophobic surfaces. Convective motion within droplets was visualized and quantified by using a high speed camera. By altering the functionality of superhydrophobic surface, two reactions were studied. In one system, singlet oxygen was generated by illuminating sensitizer particles that were partially embedded into the superhydrophobic surface. Singlet oxygen generated at the solid-liquid interface was trapped by anthracene dipropanate dianion and measured using UV spectroscopy. In a second system, the protein binding interaction between NeutrAvidin (dissolved in the droplet) and biotin (bound to the surface) was quantified by using fluorescently labeled NeutrAvidin as a probe. The binding profile was plotted as a function of time, and the kinetic rate constant was calculated. By comparing the binding rate constants with and without convection, we show that the binding reaction is limited by diffusion and that convection significantly increased the reaction rate.
33. The Secret of Chia Gel - Nanoscale 3D Network Formation
Neethu Pottackal, Priyanka Das, Malick Sematah* and George John
Department of Chemistry, Center for Discovery and Innovation (CDI) and City College of New York
Chia Gels have been associated with numerous health benefits like curbing diabetes and obesity by retarding digestion rate and quenching craving for food. However, the exact mode of gelation has always been quite ambiguous; the current explanation for the gelation process has been limited to the mucilage's swelling in or coagulation of water. We deemed this notion to fall short of explaining the fundamental science behind this observed phenomenon at nanoscale level. Gel making being one of our passion and expertise, our working hypothesis is that chia seed mucilage forms extended fibrous structures in water to ultimately form a 3D network at nanoscale. We attempted probing the mechanism of gelation in different ways including microscopic techniques. Our preliminary studies entailed gelation tests, which revealed the polar nature of the mucilage. Subsequently, we systematically did microscopic studies using optical microscope and scanning electron microscope (SEM). The SEM optical micrographs revealed bundles of fibers at microscale level while the SEM micrographs revealed a 3D network of nanoscale fibers with diameters such as 50 nm, confirming our hypothesis. This breakthrough would surely revolutionize the way we envision the gelation process of Chia and other gel forming seeds and materials.
34. Transient peptide nanostructures
Charalampos Pappas and Rein V. Ulijn
Department of Pure and Applied Chemistry, University of Strathclyde and CUNY Advanced Science Research Center
Living systems are exceptionally capable of changing their structures in response to changing situations, largely through molecular assembly and dis-assembly via competing pathways under the influence of chemical fuels. Non-equilibrium, transient nanostructures, that only exist away from thermodynamic equilibrium are increasingly of interest. Herein, we report on two different ways to trigger transience. First, the use of ultrasonic waves (80 KHz), to achieve transient reorganization of supramolecular peptide nanostructures (from fibres to micelles and twisted fibres according to the peptide sequence used), which revert back to the original state when sound is switched off. The changes observed were due to an altered balance between H-bonding and π-stacking, giving rise in changes in chiral organisation of peptide building blocks. Second, we demonstrate peptide sequence dependent formation of supramolecular nanostructures based on biocatalytic assembly and hydrolysis in chemically fuelled tripeptides using chymotrypsin. We sought to achieve control of the kinetics and consequent lifetime of the nanostructures formed by chemical design, producing nanostructures with different supramolecular chirality and hydrogen bonding.
35. Dynamic Peptide Libraries for Materials Discovery
C. G. Pappas* I. R. Sasselli, R. Shafi, D. Kalafatovic and R. V. Ulijn
Department of Pure and Applied Chemistry, University of Strathclyde and CUNY Advanced Science Research Center
Living systems are exceptionally capable of changing their structure and function in response to changing situations, largely through molecular assembly and dis-assembly via catalytic pathways, where the nature of the environment dictates the outcome of the response. There is tremendous interest in developing man-made analogues of such systems, which provides insights into the workings of biology's remarkable ability for a favorable selection of species that are best adapted and configured to their environment. The rapid responses required for biological survival are achieved through catalytic amplification, which enables dynamic transition and adaption under environmental response. Dynamic processes in living systems are regulated by balancing thermodynamic and kinetic aspects. Enzymatic pathways that activate or deactivate thermodynamic and kinetic routes leading to self-assembly (or dis-assembly) has been demonstrated, offering a tool for 'bottom-up' fabrication of peptide-based nanostructures, providing self-correction, component selection and reconfiguration, leading to the discovery of adaptive nanomaterials with enhanced plasticity and functionality.
Herein, we demonstrate i) Peptide oligomerisations from simple dipeptide building blocks through thermodynamically driven biocatalytic self-assembly, identifying the most stable supramolecular component, ii) Differential component selection and amplification accompanied with structural reconfiguration, by interplaying with environmental conditions (salts, solvents,) on Dynamic Peptide Libraries (DPLs) and iii) Shape selection and control through a direct comparison between chemically and biocatalytically driven formation of different peptides.
36. A Non-Histidine containing Short Peptide Catalyst for Esterase Activity
Krystyna L. Duncana, Roberto de la Ricaa and Rein V. Ulijna,b
aDepartment of Pure and Applied Chemistry, University of Strathclyde
bCUNY Advanced Science Research Center
The ease with which enzymes efficiently and selectively catalyse chemical reactions and biological pathways has long been admired by chemists, providing inspiration for the development of novel, synthetic catalysts to mimic the natural behavior of enzymes. Enzymes possess the ability to catalyse challenging reactions, amide bond formation for example, in aqueous environments with significant rate enhancement compared to background rates. The catalytic activity of an enzyme relies on precise and highly conserved positions of the catalytic residues within a rigid scaffold structure. Therefore, it is surprising to see that the simple dipeptide Ser-His has been reported to possess hydrolytic activity in relation to ester hydrolysis, realizing that the rigid scaffold structure is not strictly necessary for catalysis to occur. This suggests that primitive proteins could have been composed of short peptides with repeating amino acid sequences. More recent examples have built upon this by providing a short peptide nanostructure, amongst other scaffolds, for the display of catalytic residues which in turn realizes organization of these moieties enhancing catalysis. These de novo catalysts which mimic the catalytic activity of an enzyme have one thing in common; they all possess histidine which is known to play a key role in the catalytic triad of numerous enzymes. As far as we are aware, no examples of short peptidic enzyme mimics have been reported in literature whereby the histidine residue is not present.
In this present study, we demonstrate the rate enhancement of ester hydrolysis by a free peptide in solution and explore the possible mechanism of action for this catalysis. Despite this peptide lacking a histidine residue in its primary structure, it was the most convincing esterase mimic from a range of peptide sequences identified via biocatalytic screening of a phage display library. Instead, this peptide contains two cysteine residues which are known to have catalytic activity, making it highly likely that these groups play key roles in the catalytic mechanism for this peptide. Finally, we delve into the ability of this peptide to catalyse reactions in harsh conditions including pH, temperature and solvent as well as further investigating substrate scope.
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