Department of Molecular Genetics and Microbiology; Department of Microbiology and Physiological Systems
Biochemistry, Biophysics, and Structural Biology | Microbiology | Physiology
The determinants of transmembrane protein insertion orientation at the endoplasmic reticulum have been investigated in Saccharomyces cerevisiae using variants of a Type III (naturally exofacial N terminus (Nexo)) transmembrane fusion protein derived from the N terminus of Ste2p, the alpha-factor receptor. Small positive and negative charges adjacent to the transmembrane segment had equal and opposite effects on orientation, and this effect was independent of N- or C-terminal location, consistent with a purely electrostatic interaction with response mechanisms. A 3:1 bias toward Nexo insertion, observed in the absence of a charge difference, was shown to reflect the Nexo bias conferred by longer transmembrane segments. Orientation correlated best with total hydrophobicity rather than length, but it was also strongly affected by the distribution of hydrophobicity within the transmembrane segment. The most hydrophobic terminus was preferentially translocated. Insertion orientation thus depends on integration of responses to at least three parameters: charge difference across a transmembrane segment, its total hydrophobicity, and its hydrophobicity gradient. Relative signal strengths were estimated, and consequences for topology prediction are discussed. Responses to transmembrane sequence may depend on protein-translocon interactions, but responses to charge difference may be mediated by the electrostatic field provided by anionic phospholipids.
transmembrane protein insertion, yeast, endoplasmic reticulum
Rights and Permissions
© 1998 by The American Society for Biochemistry and Molecular Biology, Inc. Publisher PDF posted after 12 months as allowed by the publisher's author rights policy at http://www.jbc.org/site/misc/edpolicy.xhtml#copyright.
DOI of Published Version
J Biol Chem. 1998 Sep 18;273(38):24963-71. doi: 10.1074/jbc.273.38.24963. Link to article on publisher's site
The Journal of biological chemistry
Harley, Carol A.; Holt, Jonathan A.; Turner, Rhiannon; and Tipper, Donald J., "Transmembrane protein insertion orientation in yeast depends on the charge difference across transmembrane segments, their total hydrophobicity, and its distribution" (1998). Microbiology and Physiological Systems Publications and Presentations. 67.