Molecular Characterization of PauR and Its Role in Control of Putrescine and Cadaverine Catabolism through the  -Glutamylation Pathway in Pseudomonas aeruginosa PAO1

Abstract

bPseudomonas aeruginosa PAO1 grows on a variety of polyamines as the sole source of carbon and nitrogen. Catabolism of polyamines is mediated by the ␥-glutamylation pathway, which is complicated by the existence of multiple homologous enzymes with redundant specificities toward different polyamines for a more diverse metabolic capacity in this organism. Through a series of markerless gene knockout mutants and complementation tests, specific combinations of pauABCD (polyamine utilization) genes were deciphered for catabolism of different polyamines. Among six pauA genes, expression of pauA1, pauA2, pauA4, and pauA5 was found to be inducible by diamines putrescine (PUT) and cadaverine (CAD) but not by diaminopropane. Activation of these promoters was regulated by the PauR repressor, as evidenced by constitutively active promoters in the pauR mutant. The activities of these promoters were further enhanced by exogenous PUT or CAD in the mutant devoid of all six pauA genes. The recombinant PauR protein with a hexahistidine tag at its N terminus was purified, and specific bindings of PauR to the promoter regions of most pau operons were demonstrated by electromobility shift assays. Potential interactions of PUT and CAD with PauR were also suggested by chemical cross-linkage analysis with glutaraldehyde. In comparison, growth on PUT was more proficient than that on CAD, and this observed growth phenotype was reflected in a strong catabolite repression of pauA promoter activation by CAD but was completely absent as reflected by activation by PUT. In summary, this study clearly establishes the function of PauR in control of pau promoters in response to PUT and CAD for their catabolism through the ␥-glutamylation pathway.

Molecular characterization of lysR-lysXE, gcdR-gcdHG and amaR-amaAB operons for lysine export and catabolism: a comprehensive lysine catabolic network in Pseudomonas aeruginosa PAO1

“…Some genes that passed the initial screening parameters have been characterized in our previous studies as members of the c-glutamylation pathway for CAD and polyamine catabolism (Chou et al, 2013;Yao et al, 2011), and hence were intentionally omitted from the list. Therefore, in this table we mainly focused on genes that are either new members or less characterized members in the Lys metabolic network.…”

“…Expression of the lysine decarboxylase LdcA was found to be inducible by the arginine regulator ArgR and L-arginine but not L-lysine in P. aeruginosa PAO1 (Chou et al, 2010), making this initial step in the proposed pathway a bottleneck for L-lysine catabolism in this organism. Cadaverine, the product of LdcA, is further degraded to 5-aminovalerate through the c-glutamylation pathway for polyamine catabolism that is controlled by PauR (Chou et al, 2013;Yao et al, 2011). Both monooxygenase and decarboxylase pathways converge at 5-aminovalerate, which is then converted to glutarate by a pair of transaminase and semialdehyde dehydrogenase encoded by the gabDT operon (Chou et al, 2013;Revelles et al, 2004).…”

“…Cadaverine, the product of LdcA, is further degraded to 5-aminovalerate through the c-glutamylation pathway for polyamine catabolism that is controlled by PauR (Chou et al, 2013;Yao et al, 2011). Both monooxygenase and decarboxylase pathways converge at 5-aminovalerate, which is then converted to glutarate by a pair of transaminase and semialdehyde dehydrogenase encoded by the gabDT operon (Chou et al, 2013;Revelles et al, 2004). Glutarate is proposed to produce acetyl-CoA that can enter the Krebs cycle (Numa et al, 1964), and glutaryl-CoA dehydrogenase enzyme (an enzyme in glutarate catabolism) activity has been reported in crude extracts of P. aeruginosa (Fothergill & Guest, 1977).…”

Catabolism of biogenic amines in Pseudomonas species

“…Of these, spermidine and spermine are nucleic acid stabilizers and regulate innate immunity; they also participate in the control of inflammatory processes, in the promotion of antifungal activity and in bacterial pathogenesis/virulence (Wang et al ., ; Shah and Swiatlo, ; Yao et al ., ; Zhao et al ., ). Putrescine and cadaverine, the most abundant BPA in microbes (Chou et al ., ), are also found in different foods (meat, fish and their derivatives) and their concentrations are often used as a good indicator of foods spoilage or quality. Thus, the unpleasant odour of decomposing fish is due to the presence of putrescine, while the smell due to putrefaction of the corpses is mainly due to the presence of cadaverine.…”

“…In E. coli , some BPA (diaminopropane, putrescine, cadaverine, spermidine and spermine) are degraded through a special route which requires, in the first step, the sterification of these amines with the γ‐carboxy group of l ‐glutamate (γ‐glutamylation reaction; Kurihara et al ., , ). More recently, this pathway has also been reported in P. aeruginosa PAO1 (Chou et al ., , ; Yao et al ., ). However, in pseudomonads, this route is much more complex than in E. coli , because many different enzymes (or isozymes) can catalyse the same step and, in addition, do so with different effectiveness.…”