Dataset Information

Covalent Immobilization of Bacillus licheniformis ?-Glutamyl Transpeptidase on Aldehyde-Functionalized Magnetic Nanoparticles.

ABSTRACT: This work presents the synthesis and use of surface-modified iron oxide nanoparticles for the covalent immobilization of Bacillus licheniformis ?-glutamyl transpeptidase (BlGGT). Magnetic nanoparticles were prepared by an alkaline solution of divalent and trivalent iron ions, and they were subsequently treated with 3-aminopropyltriethoxysilane (APES) to obtain the aminosilane-coated nanoparticles. The functional group on the particle surface and the amino group of BlGGT was then cross-linked using glutaraldehyde as the coupling reagent. The loading capacity of the prepared nanoparticles for BlGGT was 34.2 mg/g support, corresponding to 52.4% recovery of the initial activity. Monographs of transmission electron microscopy revealed that the synthesized nanoparticles had a mean diameter of 15.1 ± 3.7 nm, and the covalent cross-linking of the enzyme did not significantly change their particle size. Fourier transform infrared spectroscopy confirmed the immobilization of BlGGT on the magnetic nanoparticles. The chemical and kinetic behaviors of immobilized BlGGT are mostly consistent with those of the free enzyme. The immobilized enzyme could be recycled ten times with 36.2% retention of the initial activity and had a comparable stability respective to free enzyme during the storage period of 30 days. Collectively, the straightforward synthesis of aldehyde-functionalized nanoparticles and the efficiency of enzyme immobilization offer wide perspectives for the practical use of surface-bound BlGGT.

SUBMITTER: Chen YY

PROVIDER: S-EPMC3634462 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Covalent Immobilization of Bacillus licheniformis γ-Glutamyl Transpeptidase on Aldehyde-Functionalized Magnetic Nanoparticles.

Chen Yi-Yu YY Tsai Ming-Gen MG Chi Meng-Chun MC Wang Tzu-Fan TF Lin Long-Liu LL

International journal of molecular sciences 20130226 3

This work presents the synthesis and use of surface-modified iron oxide nanoparticles for the covalent immobilization of Bacillus licheniformis γ-glutamyl transpeptidase (BlGGT). Magnetic nanoparticles were prepared by an alkaline solution of divalent and trivalent iron ions, and they were subsequently treated with 3-aminopropyltriethoxysilane (APES) to obtain the aminosilane-coated nanoparticles. The functional group on the particle surface and the amino group of BlGGT was then cross-linked usi ...[more]

PMID: 23443161

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Project description:Backgroundγ-glutamyltranspeptidase (GGT) is a bi-substrate enzyme conserved in all three domains of life. It catalyzes the cleavage and transfer of γ-glutamyl moiety of glutathione to either water (hydrolysis) or substrates like peptides (transpeptidation). GGTs exhibit great variability in their enzyme kinetics although the mechanism of catalysis is conserved. Recently, GGT has been shown to be a virulence factor in microbes like Helicobacter pylori and Bacillus anthracis. In mammalian cells also, GGT inhibition prior to chemotherapy has been shown to sensitize tumors to the therapy. Therefore, lately both bacterial and eukaryotic GGTs have emerged as potential drug targets, but the efforts directed towards finding suitable inhibitors have not yielded any significant results yet. We propose that delineating the residues responsible for the functional diversity associated with these proteins could help in design of species/clade specific inhibitors.ResultsIn the present study, we have carried out phylogenetic analysis on a set of 47 GGT-like proteins to address the functional diversity. These proteins segregate into various subfamilies, forming separate clades on the tree. Sequence conservation and motif prediction studies show that even though most of the highly conserved residues have been characterized biochemically in previous studies, a significant number of novel putative sites and motifs are discovered that vary in a clade specific manner. Many of the putative sites predicted during the functional divergence type I and type II analysis, lie close to the known catalytic residues and line the walls of the substrate binding cavity, reinforcing their role in modulating the substrate specificity, catalytic rates and stability of this protein.ConclusionThe study offers interesting insights into the evolution of GGT-like proteins in pathogenic vs. non-pathogenic bacteria, archaea and eukaryotes. Our analysis delineates residues that are highly specific to each GGT subfamily. We propose that these sites not only explain the differences in stability and catalytic variability of various GGTs but can also aid in design of specific inhibitors against particular GGTs. Thus, apart from the commonly used in-silico inhibitor screening approaches, evolutionary analysis identifying the functional divergence hotspots in GGT proteins could augment the structure based drug design approaches.

Dataset Information

Covalent Immobilization of Bacillus licheniformis ?-Glutamyl Transpeptidase on Aldehyde-Functionalized Magnetic Nanoparticles.

Publications

Covalent Immobilization of Bacillus licheniformis γ-Glutamyl Transpeptidase on Aldehyde-Functionalized Magnetic Nanoparticles.

Similar Datasets

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