Purification and characterization of Archaeoglobus fulgidus shikimate 5-dehydrogenase

摘要

Shikimate 5-dehydrogenase (EC 1.1.1.25) is an important enzyme of the aromatic amino acid biosynthesis pathway. The shikimate 5-dehydrogenase (SDH) gene from the hyperthermophile Archaeoglobus fulgidus was PCR cloned and over-expressed in E. coli. The resulting recombinant enzyme with a M/sub r/ of 27,000 was purified to homogeneity. The enzyme had a specific activity of 727 U/mg at 87/spl deg/C, and exhibited K/sub m/s for shikimate and NADP/sup +/ of 0.17 /spl plusmn/ 0.03 mM and 0.19 /spl plusmn/ 0.01, respectively. At 87/spl deg/C, the half life of the SDH was 2 hours. At 60/spl deg/C and a specific activity of 104 U/mg, the half life was 17 days. The combination of high stability and activity for this archaeal SDH may make it useful for industrial chiral synthesis.