Functional identification of novel genes involved in the glutathione-independent gentisate pathway in Corynebacterium glutamicum.
Corynebacterium glutamicum used gentisate and 3-hydroxybenzoate as its sole carbon and energy source for growth. By genome-wide data mining, a gene cluster designated ncg12918-ncg12923 was proposed to encode putative proteins involved in gentisate/3-hydroxybenzoate pathway. Genes encoding gentisate 1,2-dioxygenase (ncg12920) and fumarylpyruvate hydrolase (ncg12919) were identified by cloning and expression of each gene in Escherichia coli. The gene of ncg12918 encoding a hypothetical protein (Ncg12918) was proved to be essential for gentisate-3-hydroxybenzoate assimilation. Mutant strain RES167Deltancg12918 lost the ability to grow on gentisate or 3-hydroxybenzoate, but this ability could be restored in C. glutamicum upon the complementation with pXMJ19-ncg12918. Cloning and expression of this ncg12918 gene in E. coli showed that Ncg12918 is a glutathione-independent maleylpyruvate isomerase. Upstream of ncg12920, the genes ncg12921-ncg12923 were located, which were essential for gentisate and/or 3-hydroxybenzoate catabolism. The Ncg12921 was able to up-regulate gentisate 1,2-dioxygenase, maleylpyruvate isomerase, and fumarylpyruvate hydrolase activities. The genes ncg12922 and ncg12923 were deduced to encode a gentisate transporter protein and a 3-hydroxybenzoate hydroxylase, respectively, and were essential for gentisate or 3-hydroxybenzoate assimilation. Based on the results obtained in this study, a GSH-independent gentisate pathway was proposed, and genes involved in this pathway were identified.
Shen
Xi-Hui
XH
State Key Laboratory of Microbial Resources at the Institute of Microbiology, Chinese Academy of Sciences, Beijing 100080, People's Republic of China.
Jiang
Cheng-Ying
CY
Huang
Yan
Y
Liu
Zhi-Pei
ZP
Liu
Shuang-Jiang
SJ
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D048230:Corynebacterium glutamicum / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D049308:Dioxygenases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005841:Gentisates / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism*; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism; D062385:Hydroxybenzoates / Q000378:metabolism; D005810:Multigene Family; D019745:cis-trans-Isomerases / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D005841:Gentisates; D062385:Hydroxybenzoates; C032948:3-hydroxybenzoic acid; D049308:Dioxygenases; C026247:gentisate 1,2-dioxygenase; D006867:Hydrolases; C026249:fumarylpyruvate hydrolase; D019745:cis-trans-Isomerases; C046138:maleylpyruvate isomerase; D005978:Glutathione