Orotate phosphoribosyltransferase (OPRTase) catalyzes the magnesium-dependent conversion of alpha-D-phosphoribosylpyrophosphate (PRPP) and orotate to orotidine 5'-monophosphate (OMP) and pyrophosphate. We have determined kinetic isotope effects on the reaction of OMP with pyrophosphate and with the pyrophosphate analog phosphonoacetic acid. In the latter case, full expression of the kinetic isotope effects allowed us to calculate the structure of the transition state for the pyrophosphorylytic reaction. The transition state resembles a classical oxocarbonium ion. Using the recently reported three-dimensional structures of the OPRTase-OMP (Scapin et al., 1994) and the OPRTase-PRPP complexes (Scapin et al., 1995a), we have modeled the calculated transition state structure into the active site of OPRTase. We propose a detailed chemical mechanism which is consistent with these results.

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In this kinetic study of the thermal dissociation of methane in argon, primary product H atoms were monitored directly using the sensitive ARAS detection method. Absolute H atom concentration was determined in separate calibration experiments using N20/H2 mixtures diluted in argon. Rate constants were obtained under or near low-pressure limit conditions in shock tube experiments performed in the reflected regime. The CHI mole fraction was varied between 100 and 1300 ppm, and conversions of CHI were typically 0.2% or less. The temperature ranged from 1726 to 2134 K. Rate data were analyzed at T I 1900 K using a steady-state approximation, and at higher temperatures, rate constants were derived from simulations of [HI, profiles. The results of 37 experimental determinations were fit to the following Arrhenius expression: kodiJs( T ) = 6.8 X exp(-449OOK/T) cm3 molecule-I s-l. The overall uncertainty of values derived from this expression is estimated to be about f50%. RRKM and master equation calculations were performed to evaluate the fall-off behavior of the unimolecular rate constant and to allow comparison of the present results with those of previous studies of methane dissociation. A large discrepancy is noted, and some reasons for this disagreement are discussed.