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- Title
Identification and molecular characterization of a metagenome-derived L-lysine decarboxylase gene from subtropical soil microorganisms.
- Authors
Deng, Jie; Gao, Hua; Gao, Zhen; Zhao, Huaxian; Yang, Ying; Wu, Qiaofen; Wu, Bo; Jiang, Chengjian
- 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.
- Subjects
LYSINE decarboxylase; BIOSYNTHESIS; METAGENOMICS; MICROBIAL genomics; AMINO acids
- Publication
PLoS ONE, 2017, Vol 12, Issue 9, p1
- ISSN
1932-6203
- Publication type
Article
- DOI
10.1371/journal.pone.0185060