|
type |
Journal Article |
authors |
Deng J, Gao H, Gao Z, Zhao H, Yang Y, Wu Q, Wu B, Jiang C |
title |
Identification and molecular characterization of a metagenome-derived L-lysine decarboxylase gene from subtropical soil microorganisms |
journal |
PLoS One |
Activity |
4.1.1.18 |
Family |
4.1.1.18.b |
sel |
selected |
ui |
28931053 |
year |
(2017) |
volume |
12 |
number |
9 |
pages |
e0185060 |
| |
keywords |
doi: 10.1371/journal.pone.0185060 |
abstract |
L-lysine decarboxylase (LDC, EC 4.1.1.18) is a key enzyme in the decarboxylation of L-lysine to 1,5-pentanediamine and efficiently contributes significance to biosynthetic capability. Metagenomic technology is a shortcut approach used to obtain new genes from uncultured microorganisms. In this study, a subtropical soil metagenomic library was constructed, and a putative LDC gene named ldc1E was isolated by function-based screening strategy through the indication of pH change by L-lysine decarboxylation. Amino acid sequence comparison and homology modeling indicated the close relation between Ldc1E and other putative LDCs. Multiple sequence alignment analysis revealed that Ldc1E contained a highly conserved motif Ser-X-His-Lys (Pxl), and molecular docking results showed that this motif was located in the active site and could combine with the cofactor pyridoxal 5'-phosphate. The ldc1E gene was subcloned into the pET-30a(+) vector and highly expressed in Escherichia coli BL21 (DE3) pLysS. The recombinant protein was purified to homogeneity. The maximum activity of Ldc1E occurred at pH 6.5 and 40°C using L-lysine monohydrochloride as the substrate. Recombinant Ldc1E had apparent Km, kcat, and kcat/Km values of 1.08±0.16 mM, 5.09±0.63 s-1, and 4.73×103 s-1 M-1, respectively. The specific activity of Ldc1E was 1.53±0.06 U mg-1 protein. Identifying a metagenome-derived LDC gene provided a rational reference for further gene modifications in industrial applications. |
last changed |
2017/09/26 10:23 |
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