Genomic reconstruction of Shewanella oneidensis MR-1 metabolism reveals a previously uncharacterized machinery for lactate utilization.
The ability to use lactate as a sole source of carbon and energy is one of the key metabolic signatures of Shewanellae, a diverse group of dissimilatory metal-reducing bacteria commonly found in aquatic and sedimentary environments. Nonetheless, homology searches failed to recognize orthologs of previously described bacterial d- or l-lactate oxidizing enzymes (Escherichia coli genes dld and lldD) in any of the 13 analyzed genomes of Shewanella spp. By using comparative genomic techniques, we identified a conserved chromosomal gene cluster in Shewanella oneidensis MR-1 (locus tag: SO_1522-SO_1518) containing lactate permease and candidate genes for both d- and l-lactate dehydrogenase enzymes. The predicted d-LDH gene (dld-II, SO_1521) is a distant homolog of FAD-dependent lactate dehydrogenase from yeast, whereas the predicted l-LDH is encoded by 3 genes with previously unknown functions (lldEGF, SO_1520-SO_1518). Through a combination of genetic and biochemical techniques, we experimentally confirmed the predicted physiological role of these novel genes in S. oneidensis MR-1 and carried out successful functional validation studies in Escherichia coli and Bacillus subtilis. We conclusively showed that dld-II and lldEFG encode fully functional d-and l-LDH enzymes, which catalyze the oxidation of the respective lactate stereoisomers to pyruvate. Notably, the S. oneidensis MR-1 LldEFG enzyme is a previously uncharacterized example of a multisubunit lactate oxidase. Comparative analysis of >400 bacterial species revealed the presence of LldEFG and Dld-II in a broad range of diverse species accentuating the potential importance of these previously unknown proteins in microbial metabolism.
Pinchuk
Grigory E
GE
Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA 99352, USA.
Rodionov
Dmitry A
DA
Yang
Chen
C
Li
Xiaoqing
X
Osterman
Andrei L
AL
Dervyn
Etienne
E
Geydebrekht
Oleg V
OV
Reed
Samantha B
SB
Romine
Margaret F
MF
Collart
Frank R
FR
Scott
James H
JH
Fredrickson
Jim K
JK
Beliaev
Alexander S
AS
D001412:Bacillus subtilis / Q000235:genetics; D055162:Biocatalysis; D004926:Escherichia coli / Q000235:genetics; D016680:Genome, Bacterial; D007770:L-Lactate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D007773:Lactates / Q000378:metabolism*; D020592:Shewanella / Q000235:genetics* / Q000378:metabolism; D013237:Stereoisomerism
D007773:Lactates; D007770:L-Lactate Dehydrogenase