BIOCHEMICAL SIMILARITY OF SCHIZOSACCHAROMYCES-POMBE RAS1 PROTEIN WITHRAS2 PROTEIN OF SACCHAROMYCES-CERVISIAE

Schizosaccharomyces pombe contains single ras oncogene homologue, ras1 , that functions in the signal transduction pathway conducting the cel l's mating processes. To understand the biochemical basis of yeast ras proteins, we have purified the ras1 protein and compared the major bi ochemical constants with those of RAS2 protein from Saccharomyces cere visiae and mammalian ras proteins. The purified ras1 protein showed a remarkably high K-d value for GDP binding (178 nM) and for binding wit h ATP. In contrast, the K-d value for GTP binding and the rate of GTPa se activity were 64 nM and 77 x 10(-6) s(-1) at 37 degrees C, respecti vely; both were higher than normal p21(ras) protein, but at the same l evel 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 3 0 degrees C, respectively. On the other hand, exchange rates between b ound and free nucleotides remained almost constant throughout the test ed combination of GTP and GDP, and were several-fold lower than the bi nding rate. These results suggest that the release of the guanine nucl eotide 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.