A comprehensive characterization of novel CYP-BM3 homolog (CYP-BA) from Bacillus aryabhattai.

Autor: Bhattacharya S; Department of Biotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India., Nautiyal AK; Material Resource Efficiency Division, CSIR-Indian Institute of Petroleum, Mohkampur, Dehradun, Uttarakhand 248005, India; Academy of Scientific and Innovative Research (AcSIR), CSIR-Indian Institute of Petroleum, Mohkampur, Dehradun, Uttarakhand 248005, India., Bhattacharjee R; Department of Biotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India., Padhi AK; Laboratory for Structural Bioinformatics, Center for Biosystems Dynamics Research, RIKEN, Yokohama, Kanagawa 230-0045, Japan., Junghare V; Department of Biotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India., Bhambri M; Department of Biotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India., Dasgupta D; Material Resource Efficiency Division, CSIR-Indian Institute of Petroleum, Mohkampur, Dehradun, Uttarakhand 248005, India; Academy of Scientific and Innovative Research (AcSIR), CSIR-Indian Institute of Petroleum, Mohkampur, Dehradun, Uttarakhand 248005, India., Zhang KYJ; Laboratory for Structural Bioinformatics, Center for Biosystems Dynamics Research, RIKEN, Yokohama, Kanagawa 230-0045, Japan., Ghosh D; Material Resource Efficiency Division, CSIR-Indian Institute of Petroleum, Mohkampur, Dehradun, Uttarakhand 248005, India; Academy of Scientific and Innovative Research (AcSIR), CSIR-Indian Institute of Petroleum, Mohkampur, Dehradun, Uttarakhand 248005, India., Hazra S; Department of Biotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India; Center of Nanotechnology, Indian Institute of Technology, Roorkee, Uttarakhand 247667, India. Electronic address: shazrfbt@iitr.ac.in.
Jazyk: angličtina
Zdroj: Enzyme and microbial technology [Enzyme Microb Technol] 2021 Aug; Vol. 148, pp. 109806. Date of Electronic Publication: 2021 Apr 22.
DOI: 10.1016/j.enzmictec.2021.109806
Abstrakt: Functionalizing C-H bond poses one of the most significant challenges for chemists providing them with very few substrate-specific synthetic routes. Despite being incredibly plastic in their enzymatic ability, they are confined with deficient enzymatic action and limited explicitness of the substrates. In this study, we have endeavored to characterize novel cytochrome P450 from Bacillus aryabhattai (CYP-BA), a homolog of CYP P450-BM3, by taking interdisciplinary approaches. We conducted structure and sequence comparison to understand the conservation pattern for active site residues, conserved fold, evolutionary relationships among others. Molecular dynamics simulations were performed to understand the dynamic nature and interaction with the substrates. CYP-BA was successfully cloned, purified, and characterized. The enzyme's stability toward various physicochemical parameters was evaluated by UV-vis spectroscopy and Circular Dichroism (CD) spectroscopy. Various saturated fatty acids being the natural cytochrome P450 substrates were evaluated as catalytic efficiency of substrate oxidation by CYP-BA. The binding affinity of these natural substrates was monitored against CYP-BA by isothermal titration calorimetry (ITC). The catalytic performance of CYP-BA was satisfactory enough to proceed to the next step, that is, engineering to expand the substrate range to include polycyclic aromatic hydrocarbons (PAH). This is the first evidence of cloning, purifying and characterizing a novel homolog of CYP-BM3 to enable a better understanding of this novel biocatalyst and to provide a platform toward expanding its catalytic process through enzyme engineering.
(Copyright © 2021. Published by Elsevier Inc.)
Databáze: MEDLINE
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