Properties of the catalytic domain of CDC25, a Saccharomyces cerevisiae GDP/GTP exchange factor: comparison of its activity on full-length and C-terminal truncated RAS2 proteins.
Two C-terminal fragments (334 and 509 amino acid residues) of CDC25, a Saccharomyces cerevisiae GDP/GTP exchange factor, and the RAS2 protein were purified from E. coli, using the pGEX system. With this method it was possible to avoid in part the proteolytic phenomena that usually convert full-length RAS2 (42kDa) into 37 and 30kDa forms. Of the two CDC25 fragments containing the conserved catalytic domain, only CDC25-509 could enhance the guanine nucleotide exchange on RAS2. Comparison of the activities of RAS2-42/37kDa and RAS2-30kDa showed that the C-terminal region (112 residues) influences neither the intrinsic GDP/GTP exchange nor its stimulation by CDC25-509. RAS2-42/37kDa was somewhat more effective in enhancing the adenylylcyclase activity of a yeast membrane reconstituted system. CDC25-509 displayed a higher specific activity than the catalytic domains of the two CDC25-like proteins: S. cerevisiae SDC25 and mouse CDC25Mm.
Jacquet
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E
S.D.I. 61840 du C.N.R.S., Laboratoire de Biochimie, Ecole Polytechnique, Palaiseau, France.
Parrini
M C
MC
Bernardi
A
A
Martegani
E
E
Parmeggiani
A
A
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D005656:Fungal Proteins / Q000302:isolation & purification / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D011506:Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D018631:ras Proteins; D020666:ras-GRF1
C542351:CDC25 protein, S cerevisiae; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D005656:Fungal Proteins; D011506:Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D020666:ras-GRF1; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; C489015:RAS2 protein, S cerevisiae; D018631:ras Proteins; D000262:Adenylyl Cyclases