title: Alanine-scanning mutagenesis of WH2 domains of VopF reveals residues important for conferring lethality in a Saccharomyces cerevisiae model.
abstract: VopF, the type III effector molecule, has been implicated in the pathogenesis of non-O1, non-O139 strains of Vibrio cholerae. It is a protein of 530 amino acids, comprises of one formin homology 1-like (FH1-like) domain and three WASP homology 2 (WH2) domains. Previous works have demonstrated that WH2 domains are crucial for VopF function as a modulator of cellular actin homeostasis. Furthermore, domain deletion analysis also suggests that VopF variant constituted with only WH2 domain 3 is more efficient in restricting the growth of budding yeast than its congeners containing either only domain 1 or domain 2. Interestingly, a good degree of sequence diversity is present within each WH2 domain of VopF. In order to ascertain the importance of different amino acids in each WH2 domain, a systemic alanine scanning mutagenesis was employed. Using a yeast model system, the alanine derivatives of each amino acid of WH2 domain 1 and 3 of VopF were examined for growth restricting activity. Taken together, our mutagenesis results reveal the identification of critical residues of WH2 domain 1 and 3 of VopF.
authors.affiliation: Institute of Microbial Technology, Council of Scientific and Industrial Research, Molecular Biology Division, Chandigarh 160036, India.
authors.forename: Ranjana
authors.initials: R
authors.lastname: Tripathi
authors.forename: Vikas
authors.initials: V
authors.lastname: Kaithwas
authors.forename: Chetna
authors.initials: C
authors.lastname: Dureja
authors.forename: Saumya
authors.initials: S
authors.lastname: Raychaudhuri
mesh_terms: D000199:Actins / Q000235:genetics; D000409:Alanine / Q000235:genetics*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics; D006706:Homeostasis / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D016415:Sequence Alignment; D014734:Vibrio cholerae / Q000235:genetics; D051316:Wiskott-Aldrich Syndrome Protein / Q000235:genetics*
chemical_list: D000199:Actins; D000596:Amino Acids; D001426:Bacterial Proteins; D005656:Fungal Proteins; D051316:Wiskott-Aldrich Syndrome Protein; D000409:Alanine