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Journal of Bacteriology, August 2008, p. 5616-5623, Vol. 190, No. 16
0021-9193/08/$08.00+0     doi:10.1128/JB.00456-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Functional Characterization and Metal Ion Specificity of the Metal-Citrate Complex Transporter from Streptomyces coelicolor{triangledown}

Joshua J. Lensbouer, Ami Patel, Joseph P. Sirianni, and Robert P. Doyle*

Department of Chemistry, Center for Science and Technology, Syracuse University, Syracuse, New York

Received 2 April 2008/ Accepted 4 June 2008

Secondary transporters of citrate in complex with metal ions belong to the bacterial CitMHS family, about which little is known. The transport of metal-citrate complexes in Streptomyces coelicolor has been investigated. The best cofactor for citrate uptake in Streptomyces coelicolor is Fe3+, but uptake was also noted for Ca2+, Pb2+, Ba2+, and Mn2+. Uptake was not observed with the Mg2+, Ni2+, or Co2+ cofactor. The transportation of iron- and calcium-citrate makes these systems unique among the CitMHS family members reported to date. No complementary uptake akin to that observed for the CitH (Ca2+, Ba2+, Sr2+) and CitM (Mg2+, Ni2+, Mn2+, Co2+, Zn2+) systems of Bacillus subtilis was noted. Competitive experiments using EGTA confirmed that metal-citrate complex formation promoted citrate uptake. Uptake of free citrate was not observed. The open reading frame postulated as being responsible for the metal-citrate transport observed in Streptomyces coelicolor was cloned and overexpressed in Escherichia coli strains with the primary Fe3+-citrate transport system (fecABCDE) removed. Functional expression was successful, with uptake of Ca2+-citrate, Fe3+-citrate, and Pb2+-citrate observed. No free-citrate transport was observed in IPTG (isopropyl-β-D-thiogalactopyranoside)-induced or -uninduced E. coli. Metabolism of the Fe3+-citrate and Ca2+-citrate complexes, but not the Pb2+-citrate complex, was observed. Rationalization is based on the difference in metal-complex coordination upon binding of the metal by citrate.

* Corresponding author. Mailing address: 1-014 Center for Science and Technology, Department of Chemistry, Syracuse University, Syracuse, NY 13244-4100. Phone: (315) 443-3584. Fax: (315) 443-4070. E-mail: rpdoyle{at}syr.edu

{triangledown} Published ahead of print on 13 June 2008.

Journal of Bacteriology, August 2008, p. 5616-5623, Vol. 190, No. 16
0021-9193/08/$08.00+0     doi:10.1128/JB.00456-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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