Publications

• Cloning of the L-lactate dehydrogenase gene from the ruminal bacterium Selenomonas ruminantium HD4.

  Evans J.D., Martin S.A.

  A clone from a Selenomonas ruminantium HD4 Lambda ZAP II genomic library was isolated by its ability to complement the anaerobic growth deficiency of an Escherichia coli (pfl, ldh) double mutant. The 1.0-kb insert from the clone was sequenced and revealed a single open reading frame (ORF, 957-bp) ... >> More

  A clone from a Selenomonas ruminantium HD4 Lambda ZAP II genomic library was isolated by its ability to complement the anaerobic growth deficiency of an Escherichia coli (pfl, ldh) double mutant. The 1.0-kb insert from the clone was sequenced and revealed a single open reading frame (ORF, 957-bp) which was preceded by a putative Shine-Dalgarno (SD) sequence (AGGGGG). The potential SD sequence corresponded to 3' 16S rRNA sequences of various Selenomonas strains. The ORF was predicted to encode a protein of 318 amino acids with a calculated molecular mass of 34,975 Da and an isoelectric point of 5.54. In addition, the ORF contained 51 mol % G + C and this is consistent with the average G + C content (54%) of the S. ruminantium chromosome. The cloned S. ruminantium gene exhibited 59% nucleotide identity and 61% deduced amino acid similarity with L-lactate dehydrogenases (L-LDH) of Pediococcus acidilactici and Bacillus megaterium, respectively. Incorporation of the cloned S. ruminantium gene into E. coli DC1368 (pfl, ldh) restored anaerobic growth on glucose and L-LDH activity was detected in cell extracts. Because lactate accumulation within the rumen can be detrimental to animal performance, characterizing the gene(s) involved in lactate production by predominant ruminal bacteria will lead to a better understanding of lactate metabolism within the rumen. << Less

  Curr. Microbiol. 44:155-160(2002) [PubMed] [EuropePMC]

• IS981-mediated adaptive evolution recovers lactate production by ldhB transcription activation in a lactate dehydrogenase-deficient strain of Lactococcus lactis.

  Bongers R.S., Hoefnagel M.H.N., Starrenburg M.J.C., Siemerink M.A.J., Arends J.G.A., Hugenholtz J., Kleerebezem M.

  Lactococcus lactis NZ9010 in which the las operon-encoded ldh gene was replaced with an erythromycin resistance gene cassette displayed a stable phenotype when grown under aerobic conditions, and its main end products of fermentation under these conditions were acetate and acetoin. However, under ... >> More

  Lactococcus lactis NZ9010 in which the las operon-encoded ldh gene was replaced with an erythromycin resistance gene cassette displayed a stable phenotype when grown under aerobic conditions, and its main end products of fermentation under these conditions were acetate and acetoin. However, under anaerobic conditions, the growth of these cells was strongly retarded while the main end products of fermentation were acetate and ethanol. Upon prolonged subculturing of this strain under anaerobic conditions, both the growth rate and the ability to produce lactate were recovered after a variable number of generations. This recovery was shown to be due to the transcriptional activation of a silent ldhB gene coding for an Ldh protein (LdhB) with kinetic parameters different from those of the native las operon-encoded Ldh protein. Nevertheless, cells producing LdhB produced mainly lactate as the end product of fermentation. The mechanism underlying the ldhB gene activation was primarily studied in a single-colony isolate of the recovered culture, designated L. lactis NZ9015. Integration of IS981 in the upstream region of ldhB was responsible for transcription activation of the ldhB gene by generating an IS981-derived -35 promoter region at the correct spacing with a natively present -10 region. Subsequently, analysis of 10 independently isolated lactate-producing derivatives of L. lactis NZ9010 confirmed that the ldhB gene is transcribed in all of them. Moreover, characterization of the upstream region of the ldhB gene in these derivatives indicated that site-specific and directional IS981 insertion represents the predominant mechanism of the observed recovery of the ability to produce lactate. << Less

  J. Bacteriol. 185:4499-4507(2003) [PubMed] [EuropePMC]

• Cloning, nucleotide sequence, and transcriptional analysis of the Pediococcus acidilactici L-(+)-lactate dehydrogenase gene.

  Garmyn D., Ferain T., Bernard N., Hols P., Delcour J.

  Recombinant plasmids containing the Pediococcus acidilactici L-(+)-lactate dehydrogenase gene (ldhL) were isolated by complementing for growth under anaerobiosis of an Escherichia coli lactate dehydrogenase-pyruvate formate lyase double mutant. The nucleotide sequence of the ldhL gene predicted a ... >> More

  Recombinant plasmids containing the Pediococcus acidilactici L-(+)-lactate dehydrogenase gene (ldhL) were isolated by complementing for growth under anaerobiosis of an Escherichia coli lactate dehydrogenase-pyruvate formate lyase double mutant. The nucleotide sequence of the ldhL gene predicted a protein of 323 amino acids showing significant similarity with other bacterial L-(+)-lactate dehydrogenases and especially with that of Lactobacillus plantarum. The ldhL transcription start points in P. acidilactici were defined by primer extension, and the promoter sequence was identified as TCAAT-(17 bp)-TATAAT. This sequence is closely related to the consensus sequence of vegetative promoters from gram-positive bacteria as well as from E. coli. Northern analysis of P. acidilactici RNA showed a 1.1-kb ldhL transcript whose abundance is growth rate regulated. These data, together with the presence of a putative rho-independent transcriptional terminator, suggest that ldhL is expressed as a monocistronic transcript in P. acidilactici. << Less

  Appl. Environ. Microbiol. 61:266-272(1995) [PubMed] [EuropePMC]

• Crystal structure of Plasmodium berghei lactate dehydrogenase indicates the unique structural differences of these enzymes are shared across the Plasmodium genus.

  Winter V.J., Cameron A., Tranter R., Sessions R.B., Brady R.L.

  As Plasmodium rely extensively on homolactic fermentation for energy production, Plasmodium falciparum lactate dehydrogenase (PfLDH)--the key enzyme in this process--has previously been suggested as a novel target for antimalarials. This enzyme has distinctive kinetic and structural properties tha ... >> More

  As Plasmodium rely extensively on homolactic fermentation for energy production, Plasmodium falciparum lactate dehydrogenase (PfLDH)--the key enzyme in this process--has previously been suggested as a novel target for antimalarials. This enzyme has distinctive kinetic and structural properties that distinguish it from its human homologues. In this study, we now describe the expression, kinetic characterisation and crystal structure determination of the LDH from Plasmodium berghei. This enzyme is seen to have a similar kinetic profile to its P. falciparum counterpart, exhibiting the characteristic lack of substrate inhibition that distinguishes plasmodial from human LDHs. The crystal structure of P. berghei lactate dehydrogenase (PbLDH) shows a very similar active site arrangement to the P. falciparum enzyme. In particular, an insertion of five amino acid residues in the active site loop creates an enlarged volume in the substrate binding site, and characteristic changes in the residues lining the NADH cofactor binding pocket result in displacement of the cofactor relative to its observed position in mammalian and all other LDH structures. These results imply the special features previously described for PfLDH may be shared across the Plasmodium genus, supporting the universal application of therapeutics targeting this enzyme. << Less

  Mol. Biochem. Parasitol. 131:1-10(2003) [PubMed] [EuropePMC]