Biomedical subjects
Publications and source records attributed to Ayush Verma.
We have used mixed monolayer protected gold clusters (MMPCs) to provide flavoenzyme model systems with a high affinity and ability to modulate cofactor reduction potential.
Cytochrome c adsorbed to anionic nanoparticles is selectively proteolyzed by trypsin, providing a mechanism for the catalytic degradation of proteins.
Nanoparticles present a versatile scaffold to target biomacromolecule surfaces via complementary interactions. This review highlights some unique features of nanoparticles that make them particularly attractive resources for biomacromolecular recognition, and displays their use in modulation of structure and function of biomacromolecules.
Positively charged trimethylammonium-functionalized mixed monolayer protected clusters (MMPCs) of different chain lengths (C(8) and C(11)) have been used to bind beta-galactosidase through complementary electrostatic interactions, resulting in complete enzyme inhibition. This inhibition can be reversed in vitro by intracellular concentrations of glutathione (GSH), the main thiol component of the cell. The restoration of activity depends on the chain length of the monolayer. The activity of enzyme bound to particles with C(8) monolayer was completely restored by intracellular concentrations (1-10 mM) of GSH; however, little or no release was observed at extracellular GSH concentrations. In contrast, no restoration was observed for enzyme bound to the C(11) particles at any of the concentrations studied. Taken together, these studies demonstrate that the GSH-mediated release of enzymes bound to MMPCs can be tuned through the structure of the monolayer, a significant tool for protein and drug delivery applications.
alpha-Helices are important structural elements in proteins. To provide a scaffold for the facial recognition of peptides, we have explored the interaction of cationic mixed monolayer protected clusters (MMPCs) with a tetra-aspartate peptide in water. In these studies, substantial enhancement of peptide helicity was observed upon addition of the MMPC. Significantly, this stabilization increased with time, demonstrating templation of the monolayer to the peptide helix.
Negatively charged carboxylate-functionalized mixed monolayer protected clusters (MMPCs) effectively bind and inhibit alpha-chymotrypsin based on complementary electrostatic surface recognition. We demonstrate that this binding can be disrupted by varying the ionic strength of the medium. Enzyme activity in the presence of MMPCs increases from 5% to 97% of native activity as salt concentration is increased from 0 to 1.5 M. Variation of ionic strengths after complete binding over 13 h results only in a modest restoration of enzymatic activity (< 35%). The conformation of chymotrypsin was characterized using circular dichroism and fluorescence spectroscopy, correlating structure with enzymatic activity. This work provides a useful insight of the electrostatic influence on protein--MMPC interactions and can be applied toward MMPC-based controlled release of proteins in vivo.
Thioalkyl and thioalkylated oligo(ethylene glycol) (OEG) ligands with chain-end functionality were used to fabricate water-soluble CdSe nanoparticle scaffolds. Surface recognition of chymotrypsin (ChT) was achieved using these functionalized nanoparticle scaffolds, with three levels of interaction demonstrated: no interaction (OEG terminated with hydroxyl group), inhibition with denaturation (carboxylate-terminated thioalkyl ligands), and inhibition with retention of structure (carboxylate-terminated OEG). The latter process was reversible upon an increase in ionic strength, with essentially complete restoration of enzymatic activity.
Anionically functionalized amphiphilic nanoparticles efficiently inhibit chymotrypsin through electrostatic binding followed by protein denaturation. We demonstrate the ability to disrupt this "irreversible" inhibition of chymotrypsin through modification of the nanoparticle surface using cationic surfactants. Up to 50% of original chymotrypsin activity is rescued upon long-chain surfactant addition. Dynamic light-scattering studies demonstrate that chymotrypsin is released from the nanoparticle surface. The conformation of the rescued chymotrypsin was characterized by fluorescence and fluorescence anisotropy, indicating that chymotrypsin regains a high degree of native structure upon surfactant addition.
Positively charged trimethylammonium-functionalized mixed monolayer protected clusters (MMPCs) bind DNA through complementary electrostatic interactions, resulting in complete inhibition of DNA transcription of T7 RNA polymerase. DNA was released from the nanoparticle by intracellular concentrations of glutathione, resulting in efficient transcription. The restoration of RNA production was dose-dependent in terms of GSH, with considerable control of the release process possible through variation in monolayer structure. This work presents a new approach to controlled release of DNA, with potential applications in the creation of transfection vectors and gene regulation systems.