title: A transmembrane serine residue in the Rot1 protein is essential for yeast cell viability.
abstract: Polar residues are present in TM (transmembrane) helices and may influence the folding or association of membrane proteins. In the present study, we use an in vivo approach to analyse the functional and structural roles for amino acids in membrane-spanning motifs using the Rot1 (reversal of Tor2 lethality 1) protein as a model. Rot1 is an essential membrane protein in Saccharomyces cerevisiae and it contains a single TM domain. An alanine insertion scanning analysis of this TM helix revealed that the integrity of the central domain is essential for protein function. We identified a critical serine residue inside the helix that plays an essential role in maintaining cell viability in S. cerevisiae. Replacement of the serine residue at position 250 with a broad variety of amino acids did not affect protein targeting and location, but completely disrupted protein function causing cell death. Interestingly, substitution of the serine residue by threonine resulted in sustained cell viability, demonstrating that the hydroxy group of the TM serine side chain plays a critical role in protein function. The results of the present study indicate that Rot1 needs the TM Ser250 to interact with other membrane components and exert its functional role, avoiding exposure of the serine hydrogen-bonding group at the lipid-exposed surface.
authors.affiliation: *Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain.
authors.forename: Carlos A
authors.initials: CA
authors.lastname: Martínez-Garay
authors.affiliation: *Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain.
authors.forename: M Angeles
authors.initials: MA
authors.lastname: Juanes
authors.affiliation: *Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain.
authors.forename: J Carlos
authors.initials: JC
authors.lastname: Igual
authors.affiliation: *Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain.
authors.forename: Ismael
authors.initials: I
authors.lastname: Mingarro
authors.affiliation: *Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain.
authors.forename: M Carmen
authors.initials: MC
authors.lastname: Bañó
mesh_terms: D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000235:genetics* / Q000502:physiology; D002470:Cell Survival / Q000502:physiology*; D008565:Membrane Proteins / Q000235:genetics* / Q000502:physiology; D018832:Molecular Chaperones / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000502:physiology; D012694:Serine / Q000235:genetics* / Q000502:physiology
chemical_list: D008565:Membrane Proteins; D018832:Molecular Chaperones; C523667:ROT1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine