Gene Cloning And Function Characteristics Of A L-lysine Decarboxylase Ldc1 From Klebsiella Sp. GXW-1

L-lysine decarboxylase(EC 4.1.1.18)is within highly specific in vivo induction of intracellular enzymes in the pyridoxal phosphate effect can be L-lys decarboxylation cadaverine cadaverine aka pentanediamine,it is"iron affinity system"main ingredient,cadaverine closing porin channel also has an indispensable role,in addition to a number of strict cadaverine or anaerobic gram-negative bacteria peptides glycan composition,in terms of cadaverine plant has anti-aging,stimulating the growth of,and with the relevant plant resistance,also has been reported that cadaverine as a"second messenger"to regulate the growth of plants,except cadaverine can affect DNA,RNA,protein biosynthesis in plants,especially in recent decades studies have found that cadaverine and adipic acid may be a certain percentage of the chemical synthesis of a new order to replace nylon nylon already on the market to solve the original 6.6 environmental problems caused by the material,it will make people quite concerned.At present domestic research on lysine decarboxylase reported uncommon,but because of its low catalytic activity does not meet the industrial production.As the object of GXW-1 in this study,lysine decarboxylase enzyme were found its optimum temperature was 40?,the optimum p H was 5.4,metal ions Cd²⁺and Sr²⁺can stimulate the activity of promotion,and Trition reagent activity has a weak activation,the enzyme specific activity was 2.52 U/mg.In this study,according to Gen Bank database Klebsiella pneumoniae lysine decarboxylase gene(ldc1)primers designed,obtained by PCR amplification lysine decarboxylase gene.And p ET-32a(+)expression vector,E.Coli DH5?as host cells,recombinant expression clones.Bioinformatics analysis showed that the recombinant plasmid contains a length of 2133 bp of the ORF may encode a polypeptide of 710 amino acids,molecular weight of about 80.1 k Da,the gene was named ldc B.The recombinant plasmid into the host cell E.coli BL21(DE3),0.05%of L-lysine and IPTG inducible recombinant gene expression ldc B.By Ni affinity chromatography technology to complete the purification of recombinant protein expression.Recombinant protein found Ldc1 optimum temperature is 40?;the optimum p H is 4.6;Na⁺,Mg²⁺,Sr²⁺and Ba²⁺to Ldc B proteins activation;Chemicals 1%SDS,Tween-80 was activated by the protease activity,the enzyme specific activity was 4.14 U/mg,K_m value was 1.74 m M,V_max value was 0.934?g/min.The research work done Klebsiella pneumoniae sp.GXW-1 related physiological and biochemical function of biologically functional advantages,the completion of the corresponding lysine decarboxylase gene coding,and its analysis of bioinformatics for future industrial production cadaverine provides new material.