Publications

• Highly stable L-lysine 6-dehydrogenase from the thermophile Geobacillus stearothermophilus isolated from a Japanese hot spring: characterization, gene cloning and sequencing, and expression.

  Heydari M., Ohshima T., Nunoura-Kominato N., Sakuraba H.

  L-Lysine dehydrogenase, which catalyzes the oxidative deamination of L-lysine in the presence of NAD, was found in the thermophilic bacterium Geobacillus stearothermophilus UTB 1103 and then purified about 3,040-fold from a crude extract of the organism by using four successive column chromatograp ... >> More

  L-Lysine dehydrogenase, which catalyzes the oxidative deamination of L-lysine in the presence of NAD, was found in the thermophilic bacterium Geobacillus stearothermophilus UTB 1103 and then purified about 3,040-fold from a crude extract of the organism by using four successive column chromatography steps. This is the first report showing the presence of a thermophilic NAD-dependent lysine dehydrogenase. The product of the enzyme catalytic activity was determined to be Delta1-piperideine-6-carboxylate, indicating that the enzyme is L-lysine 6-dehydrogenase (LysDH) (EC 1.4.1.18). The molecular mass of the purified protein was about 260 kDa, and the molecule was determined to be a homohexamer with subunit molecular mass of about 43 kDa. The optimum pH and temperature for the catalytic activity of the enzyme were about 10.1 and 70 degrees C, respectively. No activity was lost at temperatures up to 65 degrees C in the presence of 5 mM L-lysine. The enzyme was relatively selective for L-lysine as the electron donor, and either NAD or NADP could serve as the electron acceptor (NADP exhibited about 22% of the activity of NAD). The Km values for L-lysine, NAD, and NADP at 50 degrees C and pH 10.0 were 0.73, 0.088, and 0.48 mM, respectively. When the gene encoding this LysDH was cloned and overexpressed in Escherichia coli, a crude extract of the recombinant cells had about 800-fold-higher enzyme activity than the extract of G. stearothermophilus. The nucleotide sequence of the LysDH gene encoded a peptide containing 385 amino acids with a calculated molecular mass of 42,239 Da. << Less

  Appl. Environ. Microbiol. 70:937-942(2004) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.

• Occurrence of L-lysine epsilon-dehydrogenase in Agrobacterium tumefaciens.

  Misono H., Nagasaki S.

  A novel amino acid dehydrogenase catalyzing the oxidative deamination of the epsilon-amino group of l-lysine was found in the crude extract of Agrobacterium tumefaciens ICR 1660. The enzyme required NAD(+) and was specific for l-lysine. The enzyme was optimally active at about pH 9.7.

  J Bacteriol 150:398-401(1982) [PubMed] [EuropePMC]

• Properties of L-lysine epsilon-dehydrogenase from Agrobacterium tumefaciens.

  Misono H., Hashimoto H., Uehigashi H., Nagata S., Nagasaki S.

  Lysine epsilon-dehydrogenase, which has been purified to homogeneity from the extract of Agrobacterium tumefaciens ICR 1600, had a molecular weight of approximately 78,000 and consisted of two subunits identical in molecular weight (about 39,000). The enzyme showed a high substrate specificity. In ... >> More

  Lysine epsilon-dehydrogenase, which has been purified to homogeneity from the extract of Agrobacterium tumefaciens ICR 1600, had a molecular weight of approximately 78,000 and consisted of two subunits identical in molecular weight (about 39,000). The enzyme showed a high substrate specificity. In addition to L-lysine, S-(beta-aminoethyl)-L-cysteine was deaminated by the enzyme, but to a far lesser extent. NAD+ and some NAD+ analogs (deamino-NAD+ and 3-acetylpyridine-NAD+) served as a cofactor. The pH optimum was at about 9.7 for the deamination of L-lysine. Although the NAD+ saturation curve was hyperbolic, a sigmoid saturation curve for L-lysine was obtained with the diluted enzyme solution, in which the dimeric enzyme was predominant. The reversible association of the enzyme to the tetramer was induced either by increasing the enzyme concentration or by addition of L-lysine. The preincubation of the enzyme with 5 mM L-lysine resulted in a 2-fold increase in the activity and gave a hyperbolic saturation curve for L-lysine. Upon modification of SH groups of the enzyme with DTNB, neither the interconversion between the dimer and the tetramer nor the activation by L-lysine occurred. These results indicated that the dimeric enzyme was activated by L-lysine and the activation resulted from the association of two dimeric enzymes to form a tetramer. << Less

  J Biochem 105:1002-1008(1989) [PubMed] [EuropePMC]

• First crystal structure of L-lysine 6-dehydrogenase as an NAD-dependent amine dehydrogenase.

  Yoneda K., Fukuda J., Sakuraba H., Ohshima T.

  A gene encoding an L-lysine dehydrogenase was identified in the hyperthermophilic archaeon Pyrococcus horikoshii. The gene was overexpressed in Escherichia coli, and its product was purified and characterized. The expressed enzyme is the most thermostable L-lysine dehydrogenase yet described, with ... >> More

  A gene encoding an L-lysine dehydrogenase was identified in the hyperthermophilic archaeon Pyrococcus horikoshii. The gene was overexpressed in Escherichia coli, and its product was purified and characterized. The expressed enzyme is the most thermostable L-lysine dehydrogenase yet described, with a half-life of 180 min at 100 degrees C. The product of the enzyme's catalytic activity is Delta(1)-piperideine-6-carboxylate, which makes this enzyme an L-lysine 6-dehydrogenase (EC 1.4.1.18) that catalyzes the reductive deamination of the epsilon-amino group and a type of NAD-dependent amine dehydrogenase. The three-dimensional structure of the enzyme was determined using the mercury-based multiple-wavelength anomalous dispersion method at a resolution of 2.44 A in the presence of NAD and sulfate ion. The asymmetric unit consisted of two subunits, and a crystallographic 2-fold axis generated the functional dimer. Each monomer consisted of a Rossmann fold domain and a C-terminal catalytic domain, and the fold of the catalytic domain showed similarity to that of saccharopine reductase. Notably, the structures of subunits A and B differed significantly. In subunit A, the active site contained a sulfate ion that was not seen in subunit B. Consequently, subunit A adopted a closed conformation, whereas subunit B adopted an open one. In each subunit, one NAD molecule was bound to the active site in an anti-conformation, indicating that the enzyme makes use of pro-R-specific hydride transfer between the two hydrides at C-4 of NADH (type A specificity). This is the first description of the three-dimensional structure of l-lysine 6-dehydrogenase as an NAD-dependent amine dehydrogenase. << Less

  J Biol Chem 285:8444-8453(2010) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.