Cooperative self-assembly of peptide gelators and proteins

Molecular self-assembly provides a versatile route for the production of nanoscale materials for medical and technological applications. Herein, we demonstrate that the cooperative self-assembly of amphiphilic small molecules and proteins can have drastic effects on supramolecular nanostructuring of resulting materials. We report that mesoscale, fractal-like clusters of proteins form at concentrations that are orders of magnitude lower compared to those usually associated with molecular crowding at room temperature. These protein clusters have pronounced effects on the molecular self-assembly of aromatic peptide amphiphiles (fluorenylmethoxycarbonyl- dipeptides), resulting in a reversal of chiral organization and enhanced order through templating and binding. Moreover, the morphological and mechanical properties of the resultant nanostructured gels can be controlled by the cooperative self-assembly of peptides and protein fractal clusters, having implications for biomedical applications where proteins and peptides are both present. In addition, fundamental insights into cooperative interplay of molecular interactions and confinement by clusters of chiral macromolecules is relevant to gaining understanding of the molecular mechanisms of relevance to the origin of life and development of synthetic mimics of living systems

Nitrate reductase is required for sclerotial development and virulence of Sclerotinia sclerotiorum

Sclerotinia sclerotiorum, the causal agent of Sclerotinia stem rot (SSR) on more than 450 plant species, is a notorious fungal pathogen. Nitrate reductase (NR) is required for nitrate assimilation that mediates the reduction of nitrate to nitrite and is the major enzymatic source for NO production in fungi. To explore the possible effects of nitrate reductase SsNR on the development, stress response, and virulence of S. sclerotiorum, RNA interference (RNAi) of SsNR was performed. The results showed that SsNR-silenced mutants showed abnormity in mycelia growth, sclerotia formation, infection cushion formation, reduced virulence on rapeseed and soybean with decreased oxalic acid production. Furthermore SsNR-silenced mutants are more sensitive to abiotic stresses such as Congo Red, SDS, H2O2, and NaCl. Importantly, the expression levels of pathogenicity-related genes SsGgt1, SsSac1, and SsSmk3 are down-regulated in SsNR-silenced mutants, while SsCyp is up-regulated. In summary, phenotypic changes in the gene silenced mutants indicate that SsNR plays important roles in the mycelia growth, sclerotia development, stress response and fungal virulence of S. sclerotiorum

Use of Activity-Based Probes to Develop High Throughput Screening Assays That Can Be Performed in Complex Cell Extracts

Background: High throughput screening (HTS) is one of the primary tools used to identify novel enzyme inhibitors. However, its applicability is generally restricted to targets that can either be expressed recombinantly or purified in large quantities. Methodology and Principal Findings: Here, we described a method to use activity-based probes (ABPs) to identify substrates that are sufficiently selective to allow HTS in complex biological samples. Because ABPs label their target enzymes through the formation of a permanent covalent bond, we can correlate labeling of target enzymes in a complex mixture with inhibition of turnover of a substrate in that same mixture. Thus, substrate specificity can be determined and substrates with sufficiently high selectivity for HTS can be identified. In this study, we demonstrate this method by using an ABP for dipeptidyl aminopeptidases to identify (Pro-Arg)2-Rhodamine as a specific substrate for DPAP1 in Plasmodium falciparum lysates and Cathepsin C in rat liver extracts. We then used this substrate to develop highly sensitive HTS assays (Z’.0.8) that are suitable for use in screening large collections of small molecules (i.e.300,000) for inhibitors of these proteases. Finally, we demonstrate that it is possible to use broad-spectrum ABPs to identify target-specific substrates. Conclusions: We believe that this approach will have value for many enzymatic systems where access to large amounts o

Control of zeolite microenvironment for propene synthesis from methanol

Optimising the balance between propene selectivity, propene/ethene ratio and catalytic stability and unravelling the explicit mechanism on formation of the first carbon–carbon bond are challenging goals of great importance in state-of-the-art methanol-to-olefin (MTO) research. We report a strategy to finely control the nature of active sites within the pores of commercial MFI-zeolites by incorporating tantalum(V) and aluminium(III) centres into the framework. The resultant TaAlS-1 zeolite exhibits simultaneously remarkable propene selectivity (51%), propene/ethene ratio (8.3) and catalytic stability (>50 h) at full methanol conversion. In situ synchrotron X-ray powder diffraction, X-ray absorption spectroscopy and inelastic neutron scattering coupled with DFT calculations reveal that the first carbon–carbon bond is formed between an activated methanol molecule and a trimethyloxonium intermediate. The unprecedented cooperativity between tantalum(V) and Brønsted acid sites creates an optimal microenvironment for efficient conversion of methanol and thus greatly promotes the application of zeolites in the sustainable manufacturing of light olefins.We thank EPSRC (EP/P011632/1), the Royal Society, National Natural Science Foundation of China (21733011, 21890761, 21673076), and the University of Manchester for funding. We thank EPSRC for funding and the EPSRC National Service for EPR Spectroscopy at Manchester. A.M.S. is supported by a Royal Society Newton International Fellowship. We are grateful to the STFC/ISIS Facility, Oak Ridge National Laboratory (ORNL) and Diamond Light Source (DLS) for access to the beamlines TOSCA/MAPS, VISION and I11/I20, respectively. We acknowledge Dr. L. Keenan for help at I20 beamline (SP23594-1). UK Catalysis Hub is kindly thanked for resources and support provided via our membership of the UK Catalysis Hub Consortium and funded by EPSRC grant: EP/R026939/1, EP/R026815/1, EP/R026645/1, EP/R027129/1 or EP/M013219/1 (biocatalysis). We acknowledge the support of The University of Manchester’s Dalton Cumbrian Facility (DCF), a partner in the National Nuclear User Facility, the EPSRC UK National Ion Beam Centre and the Henry Royce Institute. We recognise Dr. R. Edge and Dr. K. Warren for their assistance during the 60Co γ-irradiation processes. We thank Prof. A. Jentys from the Technical University of Munich for the measurement of the INS spectrum of iso-butene. We thank C. Webb, E. Enston and G. Smith for help with GC–MS; Dr. L. Hughes for help with SEM and EDX; M. Kibble for help at TOSCA/MAPS beamlines. Computing resources (time on the SCARF compute cluster for some of the CASTEP calculations) was provided by STFC’s e-Science facility. A portion of this research used resources at the Spallation Neutron Source, a DOE Office of Science User Facility operated by ORNL. The computing resources at ORNL were made available through the VirtuES and the ICE-MAN projects, funded by Laboratory Directed Research and Development programme and Compute and Data Environment for Science (CADES

Molecular hydrogels : design, synthesis, enzymatic regulation, and biological applications

In this thesis, we designed and synthesized some novel molecular hydrogelators based on short peptides or amino acids. They showed responses to environmental changes such as pH, temperature, ionic strength or catalytic reactions by enzymes. We divided the thesis into three parts. In part I, we reported the molecular hydrogels response to ligand-receptor interactions. The D-Ala-D-Ala-based hydrogels show responses to the addition of vacomycin. In part II, we talked about enzymatic regulations of molecular hydrogels. We used different enzymes (phosphatase, esterase, MMP-9, beta-lactamase) or enzymatic switch (phosphase/kinase) to control the self-assembly of small molecules in vitro or in vivo. In part III, we introduced the syntheses of novel molecular hydrogelators based on amino acids, short peptides as well as antibiotic. They formed single or multi-component hydrogels. We also reported the applications of the molecular hydrogels, including the detection of inhibitors of enzyme, removal uranium ions from the wounds on mouse, killing bacteria, etc. We have used the techniques of SEM, TEM, Rheology, Fluorescent spectra, CD spectra to characterize all the molecular hydrogels and also described the data of pHgel-sol, Tgel-sol and Cmin. in this thesis

Using enzymes to control molecular hydrogelation

The expression and distribution of enzymes differ by the types and states of cells, tissues, and organs, thus leading to diverse extracellular and intracellular environments. Using an enzymatic reaction to convert a precursor into a hydrogelator or vice versa, one can control the delivery, functions, and responses of a hydrogel according to a specific biological condition or environment, thus providing an accessible route to creating sophisticated soft materials for potential biomedical applications

Supramolecular hydrogels based on biofunctional nanofibers of self-assembled small molecules

This Feature Article summarizes the design and synthesis of supramolecular hydrogels based on nanofibers of self-assembled small bioactive molecules. The hydrogels show responses to molecular recognition and find applications in wound healing, toxin removal, and drug release. We also described our recent results of using an enzyme or enzymatic switch to trigger or regulate the self-assembly of small molecules for the generation of nanofibers and the subsequent hydrogelation. The results in this Feature Article clearly indicate that supramolecular hydrogels, as an expression of the self-assembly of molecules in water, promise a broad range of biomaterials and therapeutics

Enzymatic control of the self-assembly of small molecules: a new way to generate supramolecular hydrogels

This highlight summarizes recent advances in the enzymatic formation of supramolecular hydrogels, a new method for the creation of biofunctional soft matter. Using phosphatase, thermolysin, beta-lactamase, and phosphatase/kinase as examples, we illustrate the design and application of enzyme catalyzed or regulated formation of supramolecular hydrogels. This approach provides a new strategy to detect the presence of enzymes, screen enzyme inhibitors, assist biomineralization, assay the types of bacteria, and aid the development of smart drug delivery systems. The strategy of using enzymes to control the self-assembly of small molecules described in this highlight will lead to the further development of new materials for biomedical applications and improve understanding of molecular self-assembly in water