We have calculated the reaction rate and kinetic isotope effects for conversion of 2-phospho-D-glycerate to phosphoenolpyruvate by yeast enolase. The potential energy surface is modeled by a combined quantum mechanical/molecular mechanical method with generalized hybrid orbitals. The dynamics calculations are carried out by semiclassical variational transition state theory with multidimensional tunneling contributions. Quantum effects are included for a 25-atom cluster consisting of the substrate and part of the protein embedded in a rigid framework consisting of the rest of the protein and water. Quantum effects are important for calculating the absolute rate constant, and variational optimization of the dynamical bottleneck location is important for calculating the kinetic isotope effects. This provides the first evidence that transition state geometries are isotope dependent for enzyme reactions.