Coexpression of pyruvate decarboxylase and alcohol dehydrogenase genes in Lactobacillus brevis.
Lactobacillus brevis ATCC367 was engineered to express pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) genes in order to increase ethanol fermentation from biomass-derived residues. First, a Gram-positive Sarcina ventriculi PDC gene (Svpdc) was introduced into L. brevis ATCC 367 to obtain L. brevis bbc03. The SvPDC was detected by immunoblot using an SvPDC oligo peptide antiserum, but no increased ethanol was detected in L. brevis bbc03. Then, an ADH gene from L. brevis (Bradh) was cloned behind the Svpdc gene that generated a pdc/adh-coupled ethanol cassette pBBC04. The pBBC04 restored anaerobic growth and conferred ethanol production of Escheirichia coli NZN111 (a fermentative defective strain incapable of growing anaerobically). Approximately 58 kDa (SvPDC) and 28 kDa (BrADH) recombinant proteins were observed in L. brevis bbc04. These results indicated that the Gram-positive ethanol production genes can be expressed in L. brevis using a Gram-positive promoter and pTRKH2 shuttle vector. This work provides evidence that expressing Gram-positive ethanol genes in pentose utilizing L. brevis will further aid manipulation of this microbe toward biomass to ethanol production.
Liu
Siqing
S
Bioproducts and Biocatalysis Research Unit, Peoria, IL, USA. siqing.liu@ars.usda.gov
Dien
Bruce S
BS
Nichols
Nancy N
NN
Bischoff
Kenneth M
KM
Hughes
Stephen R
SR
Cotta
Michael A
MA
D000426:Alcohol Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D000431:Ethanol / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000502:physiology*; D005818:Genetic Engineering; D052196:Levilactobacillus brevis / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D011767:Pyruvate Decarboxylase / Q000235:genetics / Q000378:metabolism*; D012505:Sarcina / Q000235:genetics
D000431:Ethanol; D000426:Alcohol Dehydrogenase; D011767:Pyruvate Decarboxylase