Date of Award

12-1995

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Biochemistry

Supervisor

Professor D.W. Andrews

Abstract

The α subunit of the signal recognition particle receptor (SRα) is targeted to the endoplasmic reticulum membrane by a mechanism independent of the signal recognition particle-mediated translocation pathway. The membrane binding of SRα polypeptides was examined using a cell-free translation and targeting system. A membrane binding domain consisting of the amino-terminal 140 residues of SRα was identified. This domain forms a protease-resistant folded unit, and is the major site of tight binding to the β subunit of the receptor (SRβ). SRα was also shown to be a peripheral membrane protein, suggesting that it is bound to the membrane largely via interactions with the integral membrane SRβ. Co-translational targeting of SRα was correlated with a translation pause site on the SRα mRNA. An mRNA sequence at this pause site that resembles a class of retroviral frameshift sites was shown to be necessary and sufficient for a strong translation pause. SRα polypeptides synthesized from a mutant non-pausing mRNA were impaired in co-translational membrane binding, and translation-dependent localization of polysomes synthesizing SRα. These data resolve a discrepancy in earlier reports concerning the membrane attachment of SRα. Moreover, a novel pathway for polypeptide targeting and mRNA localization is proposed, and a previously undescribed signal for translation pausing is identified. Translation pausing may be general mechanism to coordinate co-translational folding, oligomeric assembly and targeting of other proteins.


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