Isolation and characterization of a Shewanella putrefaciens MR-1 electron transport regulator etrA mutant:: Reassessment of the role of EtrA

Shewanella putrefaciens MR-I has emerged as a good model to study anaerobic respiration and electron transport-linked metal reduction. Its remarkable respiratory plasticity suggests the potential for a complex regulatory system to coordinate electron acceptor use in the absence of O-2 It had previously been suggested that EtrA (electron transport regulator A), an analog of Fur (fumarate nitrate regulator) from Escherichia coli, may regulate gene expression for anaerobic electron transport. An etrA knockout strain (ETRA-153) was isolated from MR-1 using a gene replacement strategy. Reverse transcription-PCR analysis of total RNA demonstrated the loss of the etrA mRNA in ETRA-153. ETRA-153 cells retained the ability to grow on all electron acceptors tested, including fumarate, trimethylamine N-oxide (TMAO), thiosulfate, dimethyl sulfoxide, ferric citrate, nitrate, and O-2, as well as the ability to reduce ferric citrate, manganese(IV), nitrate, and nitrite. EtrA is therefore not necessary for growth on, or the reduction of, these electron acceptors. However, ETRA-153 had reduced initial growth rates on fumarate and nitrate but not on TMAO. The activities for fumarate and nitrate reductase were lower in ETRA-153, as were the levels of fumarate reductase protein and transcript. ETRA-153 was also deficient in one type of ubiquinone. These results are in contrast to those previously reported for the putative etrA mutant METR-1. Molecular analysis of METR-1 indicated that its etrA gene is not interrupted; its reported phenotype was likely due to the use of inappropriate anaerobic growth conditions.