title: Higher thermostability of l-lactate dehydrogenases is a key factor in decreasing the optical purity of d-lactic acid produced from Lactobacillus coryniformis.
abstract: Lactobacillus coryniformis is known to produce d-lactic acid as a dominant fermentation product at a cultivation temperature of approximately 30°C. However, the considerable production of l-lactic acid is observed when the fermentation temperature is greater than 40°C. Because optically pure lactates are synthesized from pyruvate by the catalysis of chiral-specific d- or l-lactate dehydrogenase, the higher thermostability of l-LDHs is assumed to be one of the key factors decreasing the optical purity of d-lactic acid produced from L. coryniformis at high temperature. To verify this hypothesis, two types of d-ldh genes and six types of l-ldh genes based on the genomic information of L. coryniformis were synthesized and expressed in Escherichia coli. Among the LDHs tested, five LDHs showed activity and were used to construct polyclonal antibodies. d-LDH1, l-LDH2, and l-LDH3 were found to be expressed in L. coryniformis by Western blotting analysis. The half-life values (t1/2) of the LDHs at 40°C were estimated to be 10.50, 41.76, and 2311min, and the T50(10) values were 39.50, 39.90, and 58.60°C, respectively. In addition, the Tm values were 36.0, 41.0, and 62.4°C, respectively, which indicates that l-LDH has greater thermostability than d-LDH. The higher thermostability of l-LDHs compared with that of d-LDH1 may be a major reason why the enantiopurity of d-lactic acid is decreased at high fermentation temperatures. The key enzymes characterized will suggest a direction for the design of genetically modified lactic acid bacteria to produce optically pure d-lactic acid.
authors.affiliation: Department of Chemical Engineering, Kwangwoon University, Seoul 139-701, Republic of Korea.
authors.forename: Sol-A
authors.initials: SA
authors.lastname: Gu
authors.affiliation: Department of Chemical Engineering, Kwangwoon University, Seoul 139-701, Republic of Korea.
authors.forename: Chanha
authors.initials: C
authors.lastname: Jun
authors.affiliation: Department of Chemical Engineering, Kwangwoon University, Seoul 139-701, Republic of Korea.
authors.forename: Jeong Chan
authors.initials: JC
authors.lastname: Joo
authors.affiliation: Korea Institute of Science and Technology (KIST), Seoul 136-791, Republic of Korea.
authors.forename: Seil
authors.initials: S
authors.lastname: Kim
authors.affiliation: Korea Research Institute of Chemical Technology (KRICT), Daejeon 305-600, Republic of Korea.
authors.forename: Seung Hwan
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Department of Chemical Engineering, Kwangwoon University, Seoul 139-701, Republic of Korea. Electronic address: metalkim@kw.ac.kr.
authors.forename: Yong Hwan
authors.initials: YH
authors.lastname: Kim
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019149:Bioreactors / Q000382:microbiology; D002152:Calorimetry, Differential Scanning; D004795:Enzyme Stability; D004926:Escherichia coli / Q000378:metabolism; D005285:Fermentation; D005813:Genes, Synthetic; D006207:Half-Life; D006358:Hot Temperature; D007770:L-Lactate Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D045022:Lactate Dehydrogenases / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D019344:Lactic Acid / Q000302:isolation & purification*; D007778:Lactobacillus / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D009904:Optical Rotation; D010802:Phylogeny; D011489:Protein Denaturation; D019289:Pyruvic Acid / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D021521:Ribotyping; D013237:Stereoisomerism
chemical_list: D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D019344:Lactic Acid; D019289:Pyruvic Acid; D045022:Lactate Dehydrogenases; D007770:L-Lactate Dehydrogenase; C071195:D-lactate dehydrogenase
keywords: Enantiopurity; Lactate dehydrogenase; Lactic acid; Lactic acid bacteria