Molecular characterization,modeling and docking of CYP107CB2 from Bacillus lehensis G1, an alkaliphile |
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| Affiliation: | 1. Enzyme and Microbial Technology Research Center, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;2. Laboratory of Enzyme Technology, Institute of Bioscience, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;3. Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;4. Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;5. Department of Chemistry, Faculty of Science, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;1. Dipartimento di Scienze del Farmaco & Drug and Food Biotechnology (DFB) Center, Università del Piemonte Orientale “A. Avogadro”, largo Donegani 2, 28100 Novara, Italy;2. Istituto di Scienze delle Produzioni Alimentari (ISPA) – CNR, via Prov.le Lecce – Monteroni, 73100 Lecce, Italy;1. Department of Cardiology, Medical University Hospital Heidelberg, Im Neuenheimer Feld 410, 69120 Heidelberg, Germany;2. HCR (Heidelberg Center for Heart Rhythm Disorders), University Hospital Heidelberg, Im Neuenheimer Feld 410, 69120 Heidelberg, Germany;3. Department of Pediatric Cardiology, University Hospital Heidelberg, Im Neuenheimer Feld 430, 69120 Heidelberg, Germany;4. Centre for Organismal Studies, Heidelberg University, Im Neuenheimer Feld 230, 69120 Heidelberg, Germany;5. Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstraße 1, 69117 Heidelberg, Germany;6. Molecular Medicine Partnership Unit, EMBL and Heidelberg University, Im Neuenheimer Feld 350, 69120 Heidelberg, Germany;7. DZHK (German Center for Cardiovascular Research), partner site Heidelberg/Mannheim, University of Heidelberg, Im Neuenheimer Feld 410, 69120 Heidelberg, Germany;8. Heidelberg Research Center for Molecular Medicine (HRCMM), Im Neuenheimer Feld 350, 69120 Heidelberg, Germany;1. Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, PR China;2. Shanghai Laiyi Center for Biopharmaceuticals R&D, Shanghai 201203, PR China;3. Shanghai Collaborative Innovation Center for Biomanufacturing Technology, Shanghai 200237, PR China |
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| Abstract: | Cytochrome P450s are a superfamily of heme monooxygenases which catalyze a wide range of biochemical reactions. The reactions involve the introduction of an oxygen atom into an inactivated carbon of a compound which is essential to produce an intermediate of a hydroxylated product. The diversity of chemical reactions catalyzed by cytochrome P450s has led to their increased demand in numerous industrial and biotechnology applications. A recent study showed that a gene sequence encoding a CYP was found in the genome of Bacillus lehensis G1, and this gene shared structural similarity with the bacterial vitamin D hydroxylase (Vdh) from Pseudonocardia autotrophica. The objectives of present study was to mine, for a novel CYP from a new isolate B. lehensis G1 alkaliphile and determine the biological properties and functionalities of CYP in this bacterium. Our study employed the usage of computational methods to search for the novel CYP from CYP structural databases to identify the conserved pattern, functional domain and sequence properties of the uncharacterized CYP from B. lehensis G1. A computational homology model of the protein’s structure was generated and a docking analysis was performed to provide useful structural knowledge on the enzyme’s possible substrate and their interaction. Sequence analysis indicated that the newly identified CYP, termed CYP107CB2, contained the fingerprint heme binding sequence motif FxxGxxxCxG at position 336-345 as well as other highly conserved motifs characteristic of cytochrome P450 proteins. Using docking studies, we identified Ser-79, Leu-81, Val-231, Val-279, Val-383, Ala-232, Thr-236 and Thr-283 as important active site residues capable of stabilizing interactions with several potential substrates, including vitamin D3, 25-hydroxyvitamin D3 and 1α-hydroxyvitamin D3, in which all substrates docked proximally to the enzyme’s heme center. Biochemical analysis indicated that CYP107CB2 is a biologically active protein to produce 1α,25-dihydroxyvitamin D3 from 1α-hydroxyvitamin D3. Based on these results, we conclude that the novel CYP107CB2 identified from B. lehensis G1 is a putative vitamin D hydroxylase which is possibly capable of catalyzing the bioconversion of parental vitamin D3 to calcitriol, or related metabolic products. |
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| Keywords: | Cytochrome P450 monooxygenase Homology modeling Molecular docking |
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