title: Functional analysis of ars gene cluster of Pannonibacter indicus strain HT23(T) (DSM 23407(T)) and identification of a proline residue essential for arsenate reductase activity.
abstract: Arsenic is a naturally occurring ubiquitous highly toxic metalloid. In this study, we have identified ars gene cluster in Pannonibacter indicus strain HT23(T) (DSM 23407(T)), responsible for reduction of toxic pentavalent arsenate. The ars gene cluster is comprised of four non-overlapping open reading frames (ORFs) encoding a transcriptional regulator (ArsR), a low molecular weight protein tyrosine phosphatases (LMW-PTPase) with hypothetical function, an arsenite efflux pump (Acr3), and an arsenate reductase (ArsC). Heterologous expression of arsenic inducible ars gene cluster conferred arsenic resistance to Escherichia coli ∆ars mutant strain AW3110. The recombinant ArsC was purified and assayed. Site-directed mutagenesis was employed to ascertain the role of specific amino acids in ArsC catalysis. Pro94X (X = Ala, Arg, Cys, and His) amino acid substitutions led to enzyme inactivation. Circular dichroism spectra analysis suggested Pro94 as an essential amino acid for enzyme catalytic activity as it is indispensable for optimum protein folding in P. indicus Grx-coupled ArsC.
authors.affiliation: Department of Biotechnology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
authors.forename: Saumya
authors.initials: S
authors.lastname: Bandyopadhyay
authors.affiliation: Department of Biotechnology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India. subratkdas@hotmail.com.
authors.forename: Subrata K
authors.initials: SK
authors.lastname: Das
mesh_terms: D019943:Amino Acid Substitution; D053502:Arsenate Reductases / Q000235:genetics* / Q000378:metabolism; D001151:Arsenic / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D007700:Kinetics; D005810:Multigene Family; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D009876:Operon; D010084:Oxidation-Reduction; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D011392:Proline / Q000737:chemistry* / Q000378:metabolism; D017510:Protein Folding; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D041902:Rhodobacteraceae / Q000201:enzymology / Q000235:genetics*; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D011392:Proline; D053502:Arsenate Reductases; D017027:Protein Tyrosine Phosphatases; D001151:Arsenic
keywords: Arsenate reductase; Circular dichroism; Heterologous expression of ars gene (s); Pannonibacter indicus; Site-directed mutagenesis; ars gene cluster