The beta-subunit of the signal recognition particle receptor is a novel GTP-binding protein without intrinsic GTPase activity.
The beta-subunit of the signal recognition particle receptor (SRbeta), a member of the Ras family of small molecular weight GTPases, is involved in the targeting of nascent polypeptide chains to the protein translocation machinery in the endoplasmic reticulum membrane. We purified SRbeta from an expressing strain of Escherichia coli and investigated the properties of the isolated GTPase. We find that, unlike other Ras family GTPases, most SRbeta purifies bound to GTP, and SRbeta-bound GTP is not easily exchanged with solution GTP. SRbeta possesses no detectable GTPase activity. Although a stable interaction between SRbeta and ribosomes is observed, SRbeta is not stimulated to hydrolyze GTP when incubated with ribosomes or ribosome-nascent chains. A GTPase mutant harboring a mutation in a region predicted to be functionally important, based on observations made in related GTPases, binds GTP with faster kinetics and appears to be a less stable protein but otherwise displays similar properties to the wild-type SRbeta GTPase. Our results demonstrate that as an isolated GTPase, SRbeta functions differently from the Arf- and Ras-type GTPases that it is most closely related to by sequence.
Legate
Kyle R
KR
Department of Biochemistry, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.
Andrews
David W
DW
D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007425:Intracellular Membranes / Q000378:metabolism; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000502:physiology*; D018000:Receptors, Peptide / Q000737:chemistry* / Q000502:physiology*; D012270:Ribosomes / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D018076:DNA, Complementary; D018160:Receptors, Cytoplasmic and Nuclear; D018000:Receptors, Peptide; C036318:signal peptide receptor; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins