Functional characterization and metal ion specificity of the metal-citrate complex transporter from Streptomyces coelicolor.
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 Fe(3+), but uptake was also noted for Ca(2+), Pb(2+), Ba(2+), and Mn(2+). Uptake was not observed with the Mg(2+), Ni(2+), or Co(2+) 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 (Ca(2+), Ba(2+), Sr(2+)) and CitM (Mg(2+), Ni(2+), Mn(2+), Co(2+), Zn(2+)) 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 Fe(3+)-citrate transport system (fecABCDE) removed. Functional expression was successful, with uptake of Ca(2+)-citrate, Fe(3+)-citrate, and Pb(2+)-citrate observed. No free-citrate transport was observed in IPTG (isopropyl-beta-d-thiogalactopyranoside)-induced or -uninduced E. coli. Metabolism of the Fe(3+)-citrate and Ca(2+)-citrate complexes, but not the Pb(2+)-citrate complex, was observed. Rationalization is based on the difference in metal-complex coordination upon binding of the metal by citrate.
Lensbouer
Joshua J
JJ
Center for Science and Technology, Department of Chemistry, Syracuse University, Syracuse, NY 13244-4100, USA.
Patel
Ami
A
Sirianni
Joseph P
JP
Doyle
Robert P
RP
D001412:Bacillus subtilis / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism*; D019343:Citric Acid / Q000378:metabolism*; D003001:Cloning, Molecular; D004533:Egtazic Acid / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000378:metabolism*; D048372:Streptomyces coelicolor / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature
D002352:Carrier Proteins; D002413:Cations, Divalent; D008670:Metals; D019343:Citric Acid; D004533:Egtazic Acid