Effect of gluconic acid as a secondary carbon source on non-growing L-lysine producers cells of Corynebacterium glutamicum. Purification and properties of 6-phosphogluconate dehydrogenase

We studied the production of L-lysine in Corynebacterium glutamicum ATCC 21 543 non growing cells obtained by nutrient limitation. Statistical analysis revealed significant differences in the L-lysine titers of glucose, glucon ic acid or glucose-gluconic acid cultures. Higher L-lysine titer obtained i n batch cultures with mixed carbon sources or gluconic acid alone were foun d to be associated with a high 6-phosphogluconate dehydrogenase activity (6 PGDH. E.C.1.1.1.44). This enzyme is a pivotal enzyme within the hexose mono phosphate pathway, and thus of importance for L-lysine production. 6PGDH wa s purified and characterized. The purified enzyme migrates as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecu lar mass of 52.5 kDa. The molecular mass of the native enzyme was estimated to be 120 kDa by molecular exclusion chromatography. thus suggesting a hom odimeric structure. The amino terminal sequence shows a strong similarity ( a match of 86% of the first 20 amino acid) to the 6PGDH from other microorg anisms such as, E. coli and B. subtilis. The pl of the dimeric native enzym e and the optimum pH were 6.2 and 8.0, respectively. For the oxidative deca rboxylation of 6-phosphogluconate, K-m of 71 muM and 43 muM were obtained f or 6-phosphogluconate and NADP(+), respectively. (C) 2001 Elsevier Science Inc. All rights reserved.