Molecular characterization of LhpR in control of hydroxyproline catabolism and transport in PAO1 Free

Abstract

Utilization of hydroxy--proline (-Hyp) in requires conversion of -Hyp to-Hyp followed by the-Hyp dehydrogenase pathway; however, the molecular mechanism in control of -Hyp catabolism and transport was not clear. DNA microarray analysis revealed twelve genes in two adjacent loci that were induced by exogenous -Hyp and -Hyp. The first locus includes encoding a Hyp epimerase (LhpA) and -Hyp dehydrogenase (LhpBEF), while the second locus codes for a putative ABC transporter (LhpPMNO), a protein of unknown function (LhpH), Hyp/Pro racemase (LhpK) and two enzymes in -Hyp catabolism (LhpC and LhpG). Proximal to these two loci, encodes a transcriptional regulator of the AraC family. The importance of these genes on -Hyp catabolism was supported by growth phenotype analysis on knockout mutants. Induction of the and promoters by exogenous-Hyp and -Hyp was demonstrated by the measurement of β-galactosidase activities from promoter- fusions in PAO1, and no induction could be detected in the Δ mutant. Induction of the promoter by -Hyp was completely abolished in the double mutant devoid of two epimerases, while the induction effect of-Hyp remained unchanged. The purified His-tagged LhpR binds specifically to the promoter regions, and formation of nucleoprotein complexes is affected by the presence of -Hyp but not -Hyp. Putative LhpR binding sites were deduced from serial deletions and comparative genomic sequence analysis. In summary, expression of genes for Hyp catabolism and uptake requires the transcriptional activator LhpR and-Hyp as the signalling compound.

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2016-07-01
2024-03-28
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