Dataset Information

Activity Augmentation of Amphioxus Peptidoglycan Recognition Protein BbtPGRP3 via Fusion with a Chitin Binding Domain.

ABSTRACT: Peptidoglycan recognition proteins (PGRPs), which have been identified in most animals, are pattern recognition molecules that involve antimicrobial defense. Resulting from extraordinary expansion of innate immune genes, the amphioxus encodes many PGRPs of diverse functions. For instance, three isoforms of PGRP encoded by Branchiostoma belcheri tsingtauense, termed BbtPGRP1~3, are fused with a chitin binding domain (CBD) at the N-terminus. Here we report the 2.7 Å crystal structure of BbtPGRP3, revealing an overall structure of an N-terminal hevein-like CBD followed by a catalytic PGRP domain. Activity assays combined with site-directed mutagenesis indicated that the individual PGRP domain exhibits amidase activity towards both DAP-type and Lys-type peptidoglycans (PGNs), the former of which is favored. The N-terminal CBD not only has the chitin-binding activity, but also enables BbtPGRP3 to gain a five-fold increase of amidase activity towards the Lys-type PGNs, leading to a significantly broadened substrate spectrum. Together, we propose that modular evolution via domain shuffling combined with gene horizontal transfer makes BbtPGRP1~3 novel PGRPs of augmented catalytic activity and broad recognition spectrum.

SUBMITTER: Wang WJ

PROVIDER: S-EPMC4610682 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Activity Augmentation of Amphioxus Peptidoglycan Recognition Protein BbtPGRP3 via Fusion with a Chitin Binding Domain.

Wang Wen-Jie WJ Cheng Wang W Luo Ming M Yan Qingyu Q Yu Hong-Mei HM Li Qiong Q Cao Dong-Dong DD Huang Shengfeng S Xu Anlong A Mariuzza Roy A RA Chen Yuxing Y Zhou Cong-Zhao CZ

PloS one 20151019 10

Peptidoglycan recognition proteins (PGRPs), which have been identified in most animals, are pattern recognition molecules that involve antimicrobial defense. Resulting from extraordinary expansion of innate immune genes, the amphioxus encodes many PGRPs of diverse functions. For instance, three isoforms of PGRP encoded by Branchiostoma belcheri tsingtauense, termed BbtPGRP1~3, are fused with a chitin binding domain (CBD) at the N-terminus. Here we report the 2.7 Å crystal structure of BbtPGRP3, ...[more]

PMID: 26479246

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Project description:Enzyme immobilization is a powerful tool not only as a protective agent against harsh reaction conditions but also for the enhancement of enzyme activity, stability, reusability, and for the improvement of enzyme properties as well. Herein, immobilization of β-glucosidase from Thermotoga maritima (Tm-β-Glu) on magnetic nanoparticles (MNPs) functionalized with chitin (Ch) was investigated. This technology showed a novel thermostable chitin-binding domain (Tt-ChBD), which is more desirable in a wide range of large-scale applications. This exclusive approach was fabricated to improve the Galacto-oligosaccharide (GOS) production from a cheap and abundant by-product such as lactose through a novel green synthesis route. Additionally, SDS-PAGE, enzyme activity kinetics, transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FT-IR) revealed that among the immobilization strategies for Thermotoga maritime-β-Glucosidase thermostable chitin-binding domain (Tm-β-Glu-Tt-ChBD) on the attractive substrate; Ch-MNPs had the highest enzyme binding capacity and GOS production ratio when compared to the native enzyme. More interestingly, a magnetic separation technique was successfully employed in recycling the immobilized Tm-β-Glu for repetitive batch-wise GOS without significant loss or reduction of enzyme activity. This immobilization system displayed an operative stability status under various parameters, for instance, temperature, pH, thermal conditions, storage stabilities, and enzyme kinetics when compared with the native enzyme. Conclusively, the GOS yield and residual activity of the immobilized enzyme after the 10th cycles were 31.23% and 66%, respectively. Whereas the GOS yield from native enzyme synthesis was just 25% after 12 h in the first batch. This study recommends applying Tt-ChBD in the immobilization process of Tm-β-Glu on Ch-MNPs to produce a low-cost GOS as a new eco-friendly process besides increasing the biostability and efficiency of the immobilized enzyme.

Dataset Information

Activity Augmentation of Amphioxus Peptidoglycan Recognition Protein BbtPGRP3 via Fusion with a Chitin Binding Domain.

Publications

Activity Augmentation of Amphioxus Peptidoglycan Recognition Protein BbtPGRP3 via Fusion with a Chitin Binding Domain.

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