Characterization of a second lysine decarboxylase isolated from Escherichia coli

Abstract

We report here on the existence of a new gene for lysine decarboxylase in Escherichia coli K-12. The hybridization experiments with a cadA probe at low stringency showed that the homologous region of cadA was located in Kohara phage clone 6F5 at 4.7 min on the E. coli chromosome. We cloned the 5.0-kb HindIII fragment of this phage clone and sequenced the homologous region of cadA. This region contained a 2,139-nucleotide open reading frame encoding a 713-amino-acid protein with a calculated molecular weight of 80,589. Overexpression of the protein and determination of its N-terminal amino acid sequence defined the translational start site of this gene. The deduced amino acid sequence showed 69.4% identity to that of lysine decarboxylase encoded by cadA at 93.7 min on the E. coli chromosome. In addition, the level of lysine decarboxylase activity increased in strains carrying multiple copies of the gene. Therefore, the gene encoding this lysine decarboxylase was designated ldc. Analysis of the lysine decarboxylase activity of strains containing cadA, ldc, or cadA ldc mutations indicated that ldc was weakly expressed under various conditions but is a functional gene in E. coli.There are two types of bacterial amino acid decarboxylase, constitutive and inducible. The former type includes decarboxylases for L-ornithine, L-arginine, S-adenosyl-L-methionine, and diaminopimelic acid (27). In Escherichia coli, a previously characterized lysine decarboxylase (EC 4.1.1.18) is encoded by cadA at 93.7 min and participates in the synthesis of cadaverine from lysine. This enzyme is inducible under anaerobic conditions at pH 5.5 and by adding lysine to the culture medium (7,19,23,27). Some evidence has been presented for the existence of a second, much less active, constitutive lysine decarboxylase in E. coli (6, 30), but the data are still too incomplete to permit any definitive conclusions on the presence of this second enzyme. Igarashi et al. made several observations concurring with the suggestion that cadaverine is actually formed by ornithine decarboxylase, which would then account for the alleged constitutive lysine decarboxylase (8). The presence of constitutive lysine decarboxylase has not been detected in organisms, except for Selenomonas ruminantium, a strictly anaerobic gram-negative bacterium (12).We previously reported the overproduction of lysine by strain WC196, a lysine analog (S-aminoethyl-L-cysteine)-resistant strain of W3110 formed by N-methyl-N-nitro-N-nitrosoguanidine (NTG) mutagenesis. During the construction of strains derived from this strain, we found that a cadA deletion mutant (WC196C) could still degrade lysine to cadaverine (15a). This finding suggested the existence of another lysine decarboxylase besides cadA in E. coli.Here, we report the existence of a new lysine decarboxylase gene, designated ldc, at 4.7 min on the E. coli chromosome, and we describe the lysine decarboxylase activities of strains containing cadA, ldc, or cadA ldc mutations. MATERIALS AND METHODSBacterial strains, bacteriophag...

Formation of a Distinctive Complex between the Inducible Bacterial Lysine Decarboxylase and a Novel AAA+ ATPase

“…6B, panel 5). However, the enzymatic activity profile and stability of LdcC differ from that of LdcI (63,64). Due to the high sequence similarity between LdcC and LdcI, we expected that LdcC might also bind RavA, however, we were surprised to find that there is no significant interaction between RavA and LdcC (Fig.…”

“…LdcI and LdcC are 69% identical and 84% similar as determined using pairwise BLAST P analysis. LdcC is expressed in E. coli at very low levels, and its exact role in the cell remains unclear (63,64). Like LdcI, LdcC is also a PLP-dependent enzyme that decarboxylates lysine and possibly assembles into a decamer (63,64) (Fig.…”

Control of 5‐FOA and 5‐FU resistance by Saccharomyces cerevisiae YJL055W

“…BLASTP searches reveal homologues in Archaea, Eubacteria, fungi, and plants (but, interestingly, apparently not in animals), suggesting that at least some aspects of Yjl055Wp function are ancient in origin and have been conserved. Some of these homologues have been annotated as 'possible lysine decarboxylases', based on a PGGxGTxxE motif that is shared with Yjl055Wp but whose association with lysine decarboxylase activity does not actually seem very clear, given that several bona fide enzymes of this type lack the motif (Kikuchi et al, 1997). Most of the Yjl055Wp homologues also share with Yjl055Wp a Rossmann-fold motif (Gx 1 -2 GxxG) that is indicative of possible nucleotide-binding activity (Kleiger and Eisenberg, 2002;Kukimoto-Niino et al, 2004).…”

Engineering of Escherichia coli to facilitate efficient utilization of isomaltose and panose in industrial glucose feedstock

“…Utilization of isomaltose and panose using GlvA and GlvC in an L-lysine-producing model strain To evaluate the effects of isomaltose and panose utilization on fermentation efficiency, we introduced the plasmids to an L-lysine-producing E. coli strain WC196LC (pCABD2) (Kojima et al 1994;Kikuchi et al 1997;Doi et al 2014). The recombinant was cultivated on L-lysine production medium supplemented with glucose or glucose combined with isomaltose, panose, or maltose (as a control).…”

“…L-Lysine production using glucose, maltose, isomaltose, and panose as carbon sources The L-lysine-producing strain WC196LC harboring pCABD2 [encoding dapA24, lysC80, dapB, and ddh (Kojima et al 1994;Kikuchi et al 1997;Doi et al 2014)] was transformed with pMW219-ΔPlac-Ptac4075-glvC and pTWV229-self-glvA-Fw. The transformant was inoculated on an LB plate containing 20 mg/L streptomycin, 100 mg/L ampicillin, and 50 mg/L kanamycin and incubated at 37°C for 24 h. Colonies were scratched off, suspended in saline, and adjusted to an OD 620 of 15.…”