Dataset Information

Genetic characterization of the resorcinol catabolic pathway in Corynebacterium glutamicum.

ABSTRACT: Corynebacterium glutamicum grew on resorcinol as a sole source of carbon and energy. By genome-wide data mining, two gene clusters, designated NCgl1110-NCgl1113 and NCgl2950-NCgl2953, were proposed to encode putative proteins involved in resorcinol catabolism. Deletion of the NCgl2950-NCgl2953 gene cluster did not result in any observable phenotype changes. Disruption and complementation of each gene at NCgl1110-NCgl1113, NCgl2951, and NCgl2952 indicated that these genes were involved in resorcinol degradation. Expression of NCgl1112, NCgl1113, and NCgl2951 in Escherichia coli revealed that NCgl1113 and NCgl2951 both coded for hydroxyquinol 1,2-dioxygenases and NCgl1112 coded for maleylacetate reductases. NCgl1111 encoded a putative monooxygenase, but this putative hydroxylase was very different from previously functionally identified hydroxylases. Cloning and expression of NCgl1111 in E. coli revealed that NCgl1111 encoded a resorcinol hydroxylase that needs NADPH as a cofactor. E. coli cells containing Ncgl1111 and Ncgl1113 sequentially converted resorcinol into maleylacetate. NCgl1110 and NCgl2950 both encoded putative TetR family repressors, but only NCgl1110 was transcribed and functional. NCgl2953 encoded a putative transporter, but disruption of this gene did not affect resorcinol degradation by C. glutamicum. The function of NCgl2953 remains unclear.

SUBMITTER: Huang Y

PROVIDER: S-EPMC1636210 | biostudies-literature | 2006 Nov

REPOSITORIES: biostudies-literature

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Genetic characterization of the resorcinol catabolic pathway in Corynebacterium glutamicum.

Huang Yan Y Zhao Ke-Xin KX Shen Xi-Hui XH Chaudhry Muhammad Tausif MT Jiang Cheng-Ying CY Liu Shuang-Jiang SJ

Applied and environmental microbiology 20060908 11

Corynebacterium glutamicum grew on resorcinol as a sole source of carbon and energy. By genome-wide data mining, two gene clusters, designated NCgl1110-NCgl1113 and NCgl2950-NCgl2953, were proposed to encode putative proteins involved in resorcinol catabolism. Deletion of the NCgl2950-NCgl2953 gene cluster did not result in any observable phenotype changes. Disruption and complementation of each gene at NCgl1110-NCgl1113, NCgl2951, and NCgl2952 indicated that these genes were involved in resorci ...[more]

PMID: 16963551

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Project description:The (pp)pGpp metabolism is an important component of bacterial physiology as it is involved in various stress responses and mechanisms of cell homeostasis, e.g., the regulation of growth. However, in order to better understand the (pp)pGpp associated regulation, it is crucial to study the molecular mechanisms of (pp)pGpp metabolism. In recent years, bioinformatic analyses of the RelA/SpoT homolog (RSH) superfamily have led to the discovery of small monofunctional RSH derivatives in addition to the well-known bifunctional Rel proteins. These are also referred to as small alarmone synthetases (SASs) or small alarmone hydrolases (SAHs). In this study, the ORF cg1485 from C. glutamicum was identified as a putative SAH encoding gene, based on a high similarity of the corresponding amino acid sequence with the (pp)pGpp hydrolysis domain. The characterization of its gene product, designated as RelHCg, represents the first functional investigation of a bacterial representative of the SAH subfamily. The predicted pyrophosphohydrolase activity was demonstrated in vivo by expression in two E. coli strains, characterized by different alarmone basal levels, as well as by in vitro analysis of the purified protein. During the assay-based analysis of hydrolysis activity in relation to the three known alarmone species, both RelHCg and the bifunctional RSH enzyme RelCg were found to exhibit a pronounced substrate inhibition for alarmone concentrations of more than 0.75 mM. This characteristic of (pp)pGpp hydrolases could be an important mechanism for realizing the bistable character of the (pp)pGpp metabolism between a (pp)pGpp basal level and stress-associated alarmone production. The deletion of relHCg caused only a minor effect on growth behavior in both wild-type background and deletion mutants with deletion of (pp)pGpp synthetases. Based on this observation, the protein is probably only present or active under specific environmental conditions. The independent loss of the corresponding gene in numerous representatives of the genus Corynebacterium, which was found by bioinformatic analyses, also supports this hypothesis. Furthermore, growth analysis of all possible deletion combinations of the three active C. glutamicum RSH genes revealed interesting functional relationships which will have to be investigated in more detail in the future.

Dataset Information

Genetic characterization of the resorcinol catabolic pathway in Corynebacterium glutamicum.

Publications

Genetic characterization of the resorcinol catabolic pathway in Corynebacterium glutamicum.

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