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Characterization of the Arsenate Respiratory Reductase from Shewanella sp. Strain ANA-3

Malasarn, Davin and Keeffe, Jennifer R. and Newman, Dianne K. (2008) Characterization of the Arsenate Respiratory Reductase from Shewanella sp. Strain ANA-3. Journal of Bacteriology, 190 (1). pp. 135-142. ISSN 0021-9193. PMCID PMC2223751. doi:10.1128/JB.01110-07. https://resolver.caltech.edu/CaltechAUTHORS:MALjbact08

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Abstract

Microbial arsenate respiration contributes to the mobilization of arsenic from the solid to the soluble phase in various locales worldwide. To begin to predict the extent to which As(V) respiration impacts arsenic geochemical cycling, we characterized the expression and activity of the Shewanella sp. strain ANA-3 arsenate respiratory reductase (ARR), the key enzyme involved in this metabolism. ARR is expressed at the beginning of the exponential phase and persists throughout the stationary phase, at which point it is released from the cell. In intact cells, the enzyme localizes to the periplasm. To purify ARR, a heterologous expression system was developed in Escherichia coli. ARR requires anaerobic conditions and molybdenum for activity. ARR is a heterodimer of ~131 kDa, composed of one ArrA subunit (~95 kDa) and one ArrB subunit (~27 kDa). For ARR to be functional, the two subunits must be expressed together. Elemental analysis of pure protein indicates that one Mo atom, four S atoms associated with a bis-molybdopterin guanine dinucleotide cofactor, and four to five [4Fe-4S] are present per ARR. ARR has an apparent melting temperature of 41°C, a Km of 5 µM, and a Vmax of 11,111 µmol of As(V) reduced min–1 mg of protein–1 and shows no activity in the presence of alternative electron acceptors such as antimonite, nitrate, selenate, and sulfate. The development of a heterologous overexpression system for ARR will facilitate future structural and/or functional studies of this protein family.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1128/JB.01110-07DOIUNSPECIFIED
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2223751/PubMed CentralArticle
ORCID:
AuthorORCID
Keeffe, Jennifer R.0000-0002-5317-6398
Newman, Dianne K.0000-0003-1647-1918
Additional Information:© 2008, American Society for Microbiology. Received 13 July 2007/ Accepted 3 September 2007. Published ahead of print on 19 October 2007. We thank Stephen Mayo (Caltech) for help with the CD analyses and members of the Newman Lab for constructive discussions. The Packard Foundation and Howard Hughes Medical Institute are acknowledged for providing financial support. Supplemental material for this article may be found at http://jb.asm.org/.
Funders:
Funding AgencyGrant Number
David and Lucile Packard FoundationUNSPECIFIED
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
Issue or Number:1
PubMed Central ID:PMC2223751
DOI:10.1128/JB.01110-07
Record Number:CaltechAUTHORS:MALjbact08
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:MALjbact08
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:9439
Collection:CaltechAUTHORS
Deposited By: Archive Administrator
Deposited On:02 Jan 2008
Last Modified:08 Nov 2021 20:59

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