Initial velocity patterns and dead-end inhibition studies with oxalylglycine suggest that the addition of NADPH, keto acid, and ammonia occurs with obligatory order. For monocarboxylic keto acids, the keto acid-ammonia initial velocity pattern is equilibrium ordered because koff is much greater than Vmax. alpha-Ketoglutarate gives substrate inhibition that is uncompetitive vs. either NADPH or ammonia in the absence of NADP+, but with high NAD+ is noncompetitive vs. NADPH and uncompetitive vs. ammonia. The inhibition is partial on both slopes and intercepts, showing that ketoglutarate slows down but does not prevent NADP+ release, and that it forms a weak but kinetically competent E-ketoglutarate complex. The apparent ordered combination of NADPH and keto acid thus results from strong synergism in binding. Deuterium isotope effects, together with the substrate inhibition patterns for ketoglutarate, show that addition of amino acid and NADP+ is random, with NADP+ being released from the ternary complex more rapidly than either glutamate or norvaline. With norvaline, hydride transfer is a major rate-limiting step, while with glutamate a step preceding hydride transfer is slower than hydride transfer. The equilibrium 18O isotope effect is 1.031 +/- 0.006 (18O enriching in ketoglutarate relative to water), but no kinetic 18O isotope effect was seen.