期刊
JOURNAL OF BACTERIOLOGY
卷 186, 期 2, 页码 454-462出版社
AMER SOC MICROBIOLOGY
DOI: 10.1128/JB.186.2.454-462.2004
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资金
- NCRR NIH HHS [RR02301, P41 RR002301, S10 RR002781, S10 RR008438] Funding Source: Medline
- NIGMS NIH HHS [R01 GM062203, GM62203] Funding Source: Medline
- NATIONAL CENTER FOR RESEARCH RESOURCES [S10RR002781, S10RR008438, P41RR002301] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [R01GM062203] Funding Source: NIH RePORTER
The propionate utilization operons of several bacteria differ from each other in the occurrence of two genes, acnD and prpF, in place of or in addition to the prpD gene encoding an Fe/S-independent 2-methylcitrate dehydratase enzyme. We cloned the acnD and prpF genes from two organisms, Shewanella oneidensis and Vibrio cholerae, and found that, together, the AcnD and PrpF proteins restored the ability of a prpD mutant strain of Salmonella enterica to grow on propionate as a source of carbon and energy. However, neither acnD nor prpF alone was able to substitute for prpD. The AcnD and PrpF proteins were isolated and biochemically analyzed. The AcnD protein required reconstitution of an Fe/S cluster for activity. All detectable AcnD activity was lost after incubation with iron-chelating agents, and no AcnD activity was observed after attempted reconstitution without iron. Nuclear magnetic resonance spectroscopy and in vitro activity assay data showed that AcnD dehydrated 2-methylcitrate and citrate to 2-methyl-cis-aconitate and cis-aconitate, respectively; AcnD also hydrated cis-aconitate. However, 2-methylisocitrate and isocitrate were not substrates for AcnD, indicating that AcnD only catalyzes the first half of the aconitase-like dehydration reactions. No aconitase-like activity was found for PrpF. It is hypothesized that, in vivo, PrpF is an accessory protein required to prevent oxidative damage of the Fe/S center of active AcnD enzyme or that it may be involved in synthesis or repair of the Fe/S cluster present in AcnD.
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