Identification and characterization of Helicobacter pylori O‐acetylserine‐dependent cystathionine β‐synthase, a distinct member of the PLP‐II family

• Published in: Molecular microbiology 2019-08, Vol.112 (2), p.718-739
• Main Authors: Devi, Suneeta, Tarique, Khaja Faisal, Ali, Mohammad Farhan, Abdul Rehman, Syed Arif, Gourinath, Samudrala
• Format: Article
• Language: English
• Subjects: Bacteria; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Binding Sites; Catalytic Domain; Cystathionine - metabolism; Cystathionine beta-Synthase - chemistry; Cystathionine beta-Synthase - genetics; Cystathionine beta-Synthase - metabolism; Cysteine; Enzyme Stability; Helicobacter pylori; Helicobacter pylori - chemistry; Helicobacter pylori - enzymology; Helicobacter pylori - genetics; Homocysteine; Homocysteine - metabolism; Hydrogen-Ion Concentration; Hydrophobicity; Kinetics; L-Serine; Methionine; Methionine - metabolism; Residues; Saccharomyces cerevisiae; Serine - analogs & derivatives; Serine - metabolism; Substrate preferences; Substrate Specificity; Substrates; Sulfhydrylase; Tyrosine
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/mmi.14315

Summary O‐acetylserine sulfhydrylase (OASS) and cystathionine β‐synthase (CBS) are members of the PLP‐II family, and involved in L‐cysteine production. OASS produces L‐cysteine via a de novo pathway while CBS participates in the reverse transsulfuration pathway. O‐acetylserine‐dependent CBS (OCBS) was previously identified as a new member of the PLP‐II family, which are predominantly seen in bacteria. The bacterium Helicobacter pylori possess only one OASS (hp0107) gene and we showed that the protein coded by this gene actually functions as an OCBS and utilizes L‐homocysteine and O‐acetylserine (OAS) to produce cystathionine. HpOCBS did not show CBS activity with the substrate L‐serine and required OAS exclusively. The HpOCBS structure in complex with methionine showed a closed cleft state, explaining the initial mode of substrate binding. Sequence and structural analyses showed differences between the active sites of OCBS and CBS, and explain their different substrate preferences. We identified three hydrophobic residues near the active site of OCBS, corresponding to one serine and two tyrosine residues in CBSs. Mutational studies were performed on HpOCBS and Saccharomyces cerevisiae CBS. A ScCBS double mutant (Y158F/Y226V) did not display activity with L‐serine, indicating indispensability of these polar residues for selecting substrate L‐serine, however, did show activity with OAS. H. pylori possess only one annotated OASS gene which functions as O‐acetylserine‐dependent CBS (OCBS) and participates in reverse transsulfuration pathway of cysteine production. OCBS is a new member in PLP‐II family, which shows activity with substrate O‐acetylserine instead of serine. Structural and sequence analyses shows three crucial residues at the active site which discriminates in substrate selection between OCBS and CBS. Mutation of residues in CBS converts into OCBS.