ABSTRACT
Glutamate dehydrogenase (GDH)a key enzyme in the nitrogen metabolism pathway catalyzes the con-version between α-ketoglutarate and glutamate reversibly using NAD(P)H as a cofactor. Based on genomic stud-ies,it was concluded that SHJG_7666 was a potential GDH in Streptomyces hygroscopicus 5008(S5008),and its expression level in vivo was positively correlated with the biosynthesis of an important aminocyclol compound vali-damycin. Phylogenetic tree analysis showed that the S5008 SHJG_7666 GDH belonged to the Glu/Leu/Phe/Val dehydrogenase family,with conserved glutamate-α-ketoglutarate binding domain and the classical GXGXXG dinu-cleotide binding motif. Further homologous modeling and structural comparison revealed that SHJG_7666 con-tained conserved Lys60,Lys78and Asp120catalytic functional sites and ligand binding residues Ser36,Gly38,Gln119 and Asp166,Asn300,Ala330. Moreover,recombinant expression of SHJG_7666 in E. coli and in vitro enzyme activity demonstrated that glutamate dehydrogenase can convert ammonium salt to glutamate with pH and temperature being optimal at 7. 5 and 37 °C respectively. Enzyme activity under optimum reaction condition has Kmvalue of (25. 3 ±9. 1)μmol/L and kcatof (3 ±0. 8)×10 -5s-1for the substrate α-ketoglutarate. Results of this study further improved the catalytic activity of SHJG_7666,thus laying the foundation for the ultimate increase of vali-damycin production.
ABSTRACT
Carbon-nitrogen lyases (E.C.4.3) are a group of enzymes that release ammonia, amidine or amino group etc, and also show ability to form double bond or ring structure. Specifically, enzymes forming amino group are called amine-lyases (E.C.4.3.3), which are critical in the industrial production of many medicine intermediates. In this review is a summary of four major amine-lyases in terms of their source, enzymatic characteristics and their applications in preparation of pharmaceutical intermediates.