RRC ID 29214
Author Saito Y, Kimura K, Oka T, Nakano A.
Title Activities of mutant Sar1 proteins in guanine nucleotide binding, GTP hydrolysis, and cell-free transport from the endoplasmic reticulum to the Golgi apparatus.
Journal J Biochem
Abstract Sar1p belongs to a unique subfamily of the small GTPase superfamily and is essential for the formation of vesicles that transport proteins from the endoplasmic reticulum to the Golgi apparatus. We have obtained mutants of the yeast SAR1 gene, which show several different phenotypes in cell growth and protein transport [Nakano, A. , Otsuka, H., Yamagishi, M., Yamamoto, E., Kimura, K., Nishikawa, S., and Oka, T. (1994) J. Biochem. 116, 243-247; Yamanushi, T., Hirata, A., Oka, T., and Nakano, A. (1996) ibid. 120, 452-458]. In this study, we have purified five mutant Sar1 proteins using an Escherichia coli expression system and characterized their biochemical properties in detail. Three of them prefer GDP binding to GTP binding and are thus regarded as GDP-form mutants, and one is insensitive to the GTPase-activating protein and is almost fixed in the GTP-bound state. The GDP mutants are defective in vesicle formation in vitro, whereas the GTP mutant can drive vesicle formation but not the overall transport to the Golgi. These mutants will be useful for further understanding of the regulation of the GTPase cycle of Sar1p.
Volume 124(4)
Pages 816-23
Published 1998-10-1
DOI 10.1093/oxfordjournals.jbchem.a022185
PMID 9756629
MeSH Amino Acid Substitution Binding Sites Cell-Free System Cloning, Molecular Endoplasmic Reticulum / metabolism* Escherichia coli GTP Phosphohydrolases / metabolism GTP-Binding Proteins / genetics GTP-Binding Proteins / isolation & purification GTP-Binding Proteins / metabolism* GTPase-Activating Proteins Golgi Apparatus / metabolism* Guanosine Diphosphate / metabolism* Guanosine Triphosphate / metabolism* Kinetics Monomeric GTP-Binding Proteins* Mutagenesis, Site-Directed Point Mutation Proteins / metabolism Recombinant Proteins / isolation & purification Recombinant Proteins / metabolism Saccharomyces cerevisiae / genetics* Saccharomyces cerevisiae / metabolism* Saccharomyces cerevisiae Proteins* Vesicular Transport Proteins
IF 2.476
Times Cited 28
DNA material pMYE3-1 (RDB08876) pMYE3-2 (RDB08877) pMYE3-3 (RDB08878) pMYE3-4 (RDB08879) pMYE3-5 (RDB08880) pMYE3-6 (RDB08881) pMYE3-7 (RDB08882) pMYE3-8 (RDB08883) pMYE3-9 (RDB08884)