NMR studies of mating factors from the yeast Saccharomyces cerevisiae.
Item
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Title
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NMR studies of mating factors from the yeast Saccharomyces cerevisiae.
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Identifier
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AAI9431358
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identifier
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9431358
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Creator
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Gounarides, John Stephen.
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Contributor
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Adviser: Fred Naider
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Date
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1994
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Language
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English
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Publisher
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City University of New York.
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Subject
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Chemistry, Physical | Biophysics, General
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Abstract
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Mating in Saccharomyces cerevisiae is mediated by the {dollar}\alpha{dollar}-factor (WHWLQLKPGQPMY) and the a-factor (YIIKGVFWDPAC (S-farnesyl) OCH{dollar}\sb3{dollar}). The (scD-Ala{dollar}\sp9{dollar}) - and (scL-Ala{dollar}\sp9{dollar}) {dollar}alpha{dollar}-factors were synthesized and examined in solution and in the presence of lipid vesicles by NMR spectroscopy. NOE and NH d{dollar}\delta{dollar}/dT data indicated that residues 7-10 of the (scD-Ala{dollar}\sp9{dollar}) {dollar}\alpha{dollar}-factor adopt a Type II {dollar}\beta{dollar}-turn in DMSO and aqueous solutions. The 10-fold less active (scL-Ala{dollar}\sp9{dollar}) analogue did not adopt a regular secondary structure. Transfer NOE data suggested that the Type II {dollar}\beta{dollar}-turn conformation of the (scD-Ala{dollar}\sp9{dollar}) {dollar}\alpha{dollar}-factor is maintained in the lipid bound state.;Several cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7,{dollar} X{dollar}\sp9{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) {dollar}\alpha{dollar}-factor analogues (X = scD-Ala, scL-Ala, scD-Val, or Gly) were studied in DMSO/water (80:20) and aqueous solution using NMR spectroscopy. NOE parameters, NH d{dollar}\delta{dollar}/dT, and {dollar}\sp3J\sb{lcub}\alpha{lcub}\rm NH{rcub}{rcub}{dollar} coupling constant indicate that residues 7-10 of cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar}, scD-Ala {dollar}\sp9{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) - and cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar}, scD-Val{dollar}\sp9{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) {dollar}\alpha{dollar}-factor adopt a Type II {dollar}\beta{dollar}-turn, whereas cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar}, scL-Ala{dollar}\sp9{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) {dollar}\alpha{dollar}-factor adopts a Type I {dollar}\beta{dollar}-turn. In water cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) {dollar}\alpha{dollar}-factor adopts at least two distinct conformations, however, in DMSO/water cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) {dollar}\alpha{dollar}-factor assumes a Type II {dollar}\beta{dollar}-turn.;Vibrational circular dichroism (VCD) studies suggested that cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar}, scD-Ala{dollar}\sp9{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) - and cyclo{dollar}\sp{lcub}7,10{rcub}{dollar} (Cys{dollar}\sp7{dollar}, scL-Ala{dollar}\sp9{dollar},Cys{dollar}\sp{10}{dollar},Nle{dollar}\sp{12}{dollar}) {dollar}\alpha{dollar}-factors adopt a transient {dollar}\beta{dollar}-sheet and that VCD could be used to distinguish Type I from Type II {dollar}\beta{dollar}-turns.;{dollar}\alpha{dollar}-Factor and five {dollar}\alpha{dollar}-factor analogues were studied in DMSO-d{dollar}\sb6{dollar}. NH d{dollar}\delta{dollar}/dT, {dollar}\sp3{dollar}J{dollar}\sb{lcub}\alpha{lcub}\rm NH{rcub}{rcub}{dollar} coupling constant, and sequential NOESY data indicated that a-factor is predominantly unstructured in DMSO. Similar results were obtained for the other peptides indicating that S-prenylation of Cys{dollar}\sp{12}{dollar} does not affect the conformation of these peptides.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
