Dataset Information

Crystal structure and functional insight of HP0420-homolog from Helicobacter felis.

ABSTRACT: Helicobacter pylori infect more than half of the world's population and are considered a cause of peptic ulcer disease and gastric cancer. Recently, hypothetical gene HP0421 was identified in H. pylori as a cholesterol alpha-glucosyltransferase, which is required to synthesize cholesteryl glucosides, essential cell wall components of the bacteria. In the same gene-cluster, HP0420 was co-identified, whose function remains unknown. Here we report the crystal structure of HP0420-homolog of H. felis (HF0420) to gain insight into the function of HP0420. The crystal structure, combined with size-exclusion chromatography, reveals that HF0420 adopts a homodimeric hot-dog fold. The crystal structure suggests that HF0420 has enzymatic activity that involves a conserved histidine residue at the end of the central alpha-helix. Subsequent biochemical studies provide clues to the function of HP0420 and HF0420.

SUBMITTER: Piao S

PROVIDER: S-EPMC2891984 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Crystal structure and functional insight of HP0420-homolog from Helicobacter felis.

Piao Shunfu S Jin Xiao Ling XL Yun Bo-Young BY Kim Nahee N Cho Hyun-Soo HS Fukuda Minoru M Lee Heeseob H Ha Nam-Chul NC

Biochemical and biophysical research communications 20100317 4

Helicobacter pylori infect more than half of the world's population and are considered a cause of peptic ulcer disease and gastric cancer. Recently, hypothetical gene HP0421 was identified in H. pylori as a cholesterol alpha-glucosyltransferase, which is required to synthesize cholesteryl glucosides, essential cell wall components of the bacteria. In the same gene-cluster, HP0420 was co-identified, whose function remains unknown. Here we report the crystal structure of HP0420-homolog of H. felis ...[more]

PMID: 20302842

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Project description:Serine hydroxymethyltransferase (SHMT), encoded by the glyA gene, is a ubiquitous pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the formation of glycine from serine. The thereby generated 5,10-methylene tetrahydrofolate (MTHF) is a major source of cellular one-carbon units and a key intermediate in thymidylate biosynthesis. While in virtually all eukaryotic and many bacterial systems thymidylate synthase ThyA, SHMT and dihydrofolate reductase (DHFR) are part of the thymidylate/folate cycle, the situation is different in organisms using flavin-dependent thymidylate synthase ThyX. Here the distinct catalytic reaction directly produces tetrahydrofolate (THF) and consequently in most ThyX-containing organisms, DHFR is absent. While the resulting influence on the folate metabolism of ThyX-containing bacteria is not fully understood, the presence of ThyX may provide growth benefits under conditions where the level of reduced folate derivatives is compromised. Interestingly, the third key enzyme implicated in generation of MTHF, serine hydroxymethyltransferase (SHMT), has a universal phylogenetic distribution, but remains understudied in ThyX-containg bacteria. To obtain functional insight into these ThyX-dependent thymidylate/folate cycles, we characterized the predicted SHMT from the ThyX-containing bacterium Helicobacter pylori. Serine hydroxymethyltransferase activity was confirmed by functional genetic complementation of a glyA-inactivated E. coli strain. A H. pylori ΔglyA strain was obtained, but exhibited markedly slowed growth and had lost the virulence factor CagA. Biochemical and spectroscopic evidence indicated formation of a characteristic enzyme-PLP-glycine-folate complex and revealed unexpectedly weak binding affinity of PLP. The three-dimensional structure of the H. pylori SHMT apoprotein was determined at 2.8Ǻ resolution, suggesting a structural basis for the low affinity of the enzyme for its cofactor. Stabilization of the proposed inactive configuration using small molecules has potential to provide a specific way for inhibiting HpSHMT.

Dataset Information

Crystal structure and functional insight of HP0420-homolog from Helicobacter felis.

Publications

Crystal structure and functional insight of HP0420-homolog from Helicobacter felis.

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