Role of glycine-82 as a pivot point during the transition from the inactive to the active form of the yeast Ras2 protein

Ras proteins bind either GDP or GTP with high affinity. However, only the GTP-bound form of the yeast Ras2 protein is able to stimulate adenylyl cyclase. To identify amino acid residues that play a role in the conversion from the GDP-bound to the GTP-bound state of Ras proteins, we have searched for...

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Bibliographic Details
Published in:	FEBS letters Vol. 281; no. 1; pp. 235 - 239
Main Authors:	Kavounis, Constantin, Verrotti, Arturo C., De Vendittis, Emmanuele, Bozopoulos, Anastasios, Di Blasi, Francesco, Zahn, Regina, Crechet, Jean Bernard, Parmeggiani, Andrea, Tsernoglou, Demetrios, Fasano, Ottavio
Format:	Journal Article
Language:	English
Published:	Amsterdam Elsevier B.V 09-04-1991 Elsevier
Subjects:	[ethylenedinitrilo]tetraacetic acid Amino Acid Sequence Analytical, structural and metabolic biochemistry Binding Sites Biological and medical sciences EDTA Escherichia coli - genetics Fundamental and applied biological sciences. Psychology Fungal Proteins - genetics Fungal Proteins - isolation & purification Fungal Proteins - metabolism GDP-βS Glycine Gpp(NH)p GTP-Binding Proteins - metabolism Guanosine diphosphate Guanosine Diphosphate - metabolism Guanosine triphosphate Guanosine Triphosphate - metabolism guanosine-5′-O-(2-thiodiphosphate) guanyl-5′-yl imidodiphosphate Guanylyl Imidodiphosphate - metabolism Miscellaneous Models, Structural Mutagenesis, Site-Directed PAGE Plasmids Point mutation polyacrylamide gel electrophoresis Protein Conformation Proteins Ras ras Proteins Recombinant Proteins - isolation & purification Recombinant Proteins - metabolism Restriction Mapping S. cerevisae Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae - metabolism Saccharomyces cerevisiae Proteins SDS sodium dodecyl sulfate GDP-βS Guanosine triphosphate SDS Ras S. cerevisae Guanosine diphosphate Gpp(NH)p Point mutation PAGE EDTA
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Summary:	Ras proteins bind either GDP or GTP with high affinity. However, only the GTP-bound form of the yeast Ras2 protein is able to stimulate adenylyl cyclase. To identify amino acid residues that play a role in the conversion from the GDP-bound to the GTP-bound state of Ras proteins, we have searched for single amino acid substitutions that selectively affected the binding of one of the two nucleotides. We have found that the replacement of glycine-82 of the Ras2 protein by serine resulted in an increased rate of dissociation of Gpp(NH)p, a nonhydrolysable analog of GTP, while the GDP dissociation rate was not significantly modified. Glycine-82 resides in a region that is highly conserved between the yeast and human proteins. However, this residue is structurally distant from residues that participate in the binding of the nucleotide, as determined from the crystal structure of the human H- ras gene product. Therefore, the ability of the nucleotide binding site to discriminate between GDP and GTP is dependent not only on residues that are spatially close to the nucleotide, but also on distant amino acids. This is in agreement with the role of glycine-82 as a pivot point during the transition from the GDP- to the GTP-bound form of the Ras proteins.
Bibliography:	ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23
ISSN:	0014-5793 1873-3468
DOI:	10.1016/0014-5793(91)80401-N