Abstract
Understanding the remarkable catalytic efficiency of enzymes has been the object of intense study for decades. Single molecule fluorescence microscopy has the potential of providing fresh insight into the mechanism of action of enzymes. In particular, single molecular events can provide information about states that may be obscured with ensemble measurements and about the heterogeneity of catalysis that may be produced by the multiple protein conformations that exist. Single molecule and ensemble experiments with dihydrofolate reductase demonstrate that multiple conformations are an essential part of the catalytic reaction and that the conformational changes are propagated throughout the enzyme molecule. The coupling of conformational changes to catalysis within a single molecule can be demonstrated by site specific mutagenesis and isotope rate effects (NADPH vs. NADPD). The results for dihydrofolate reductase, as well as those obtained with single molecule methods for other enzymes, will be discussed within the context of the static and dynamic roles of the enzyme structure in catalysis. This work was supported by NIH grant GM-65128.
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