Biochemical similarity of Schizosaccharomyces pombe ras1 protein with RAS2 protein of Saccharomyces cervisiae.
Schizosaccharomyces pombe contains single ras oncogene homologue, ras1, that functions in the signal transduction pathway conducting the cell's mating processes. To understand the biochemical basis of yeast ras proteins, we have purified the ras1 protein and compared the major biochemical constants with those of RAS2 protein from Saccharomyces cerevisiae and mammalian ras proteins. The purified ras1 protein showed a remarkably high Kd value for GDP binding (178 nM) and for binding with ATP. In contrast, the Kd value for GTP binding and the rate of GTPase activity were 64 nM and 77 x 10(-6) s-1 at 37 degrees C, respectively; both were higher than normal p21ras protein, but at the same level as the RAS2 protein. We directly measured rate of GTP binding and GDP binding which were 3.9 x 10(-3) s-1 and 1.8 x 10(-3) s-1 at 30 degrees C, respectively. On the other hand, exchange rates between bound and free nucleotides remained almost constant throughout the tested combination of GTP and GDP, and were several-fold lower than the binding rate. These results suggest that the release of the guanine nucleotide is the rate-limiting step in the ras-GTP/GDP cycle. As a whole, the biochemical properties of the ras1 protein are close to those of the RAS2 protein, although these two proteins function differently in the signal transduction pathway in the cells.
D000595:Amino Acid Sequence; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006150:Guanine Nucleotides / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D012568:Schizosaccharomyces / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins; D018631:ras Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D017931:DNA Primers; D005656:Fungal Proteins; D006150:Guanine Nucleotides; D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C489014:RAS1 protein, S cerevisiae; C489015:RAS2 protein, S cerevisiae; C492657:Ras1 protein, S pombe; D018631:ras Proteins