Abstract
Cadaverine is produced in organisms from the amino acid lysine in a decarboxylation reaction catalyzed by lysine decarboxylase (EC 4.1.1.18). The inducible lysine decarboxylase CadA plays a vital role in acid stress response for enteric bacteria. Vibrio vulnificus is an extremely virulent human pathogen causing gastroenteritis when the acid conditions that prevent survival of V. vulnificus in the stomach or small intestine are overcome. A gene encoding CadA was identified from V. vulnificus. Subsequent analyses showed that CadA from V. vulnificus (VvCadA) is a decamer with a 82-kDa subunit. Homogenous VvCadA was purified from Escherichia coli and used for lysine decarboxylation with an optimal pH of 6.0 and optimal temperature of 37°C. The apparent Vmax and Km for lysine were 9.45 ± 0.24 μM/min and 0.45 ± 0.05 mM, respectively. Mutation analysis suggested that the amino-acid-binding pyridoxal phosphate, the cofactor of the enzyme, plays a vital role in the reaction. Mutation of the negatively charged residues interacting with lysine also affected the activity of the enzyme to some extent. Quantitative RT-PCR showed that expression of VvcadA was up-regulated under low pH, low salinity, and oxidative stresses. Furthermore, the concentration of cadaverine released to the cell exterior also increased under these stresses. Protein sequence similarity network (SSN) analysis indicated that lysine decarboxylases with ornithine decarboxylases and arginine decarboxylases shared a common ancestor, and that lysine decarboxylases are more conserved during evolution. Our data provide evidence for the biochemical characteristics and important roles of VvCadA under stress conditions.
Highlights
Of the diverse marine bacteria, Vibrio vulnificus is one of the most common causes of gastroenteritis and sepsis (Morris, 1988)
The VvcadA ORF contains 2136 nucleotides coding for a protein of 711 amino acids, with a theoretical pI of 5.53 and MW of 80.432
This VvcadA gene was obtained from V. vulnificus genomic DNA through PCR
Summary
Of the diverse marine bacteria, Vibrio vulnificus is one of the most common causes of gastroenteritis and sepsis (Morris, 1988). Lysine Decarboxylase in Vibrio vulnificus contribute to the disease caused by V. vulnificus (Wright et al, 1981; Smith and Merkel, 1982; Simpson and Oliver, 1983; Testa et al, 1984; Kothary and Kreger, 1987; Simpson et al, 1987; Lee et al, 2004b), and motility could be another virulence determinant of this organism (Lee et al, 2004a). Two different mechanisms are used in this acid tolerance (Jones and Oliver, 2009): up-regulation of lysine decarboxylase and manganese superoxide dismutase (MnSOD) under acid and secondary oxidative stress. Lysine decarboxylase converts L-lysine into cadaverine (1,5-pentanediamine), which is an acid neutralizer (through deamination) and superoxide radical scavenger; MnSOD is involved in acid neutralization and reduction of oxidative stress (Kim et al, 2005)
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