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. 2013 Jul 30;288(37):26350–26356. doi: 10.1074/jbc.M113.487447

FIGURE 2.

FIGURE 2.

Solvent viscosity and isotope effects reveal the rate-determining step(s) in WT and variant ssIGPS enzymes. A, the kinetic mechanism of IGPS. There is a substantial SVE when the rate-determining step is diffusion-controlled, such as in product release, similar to what occurs at low temperatures (11). A substantial SDKIE is present only when the condensation step is rate-determining (see “Experimental Procedures” and supplemental “Experimental Procedures”). B, summary of the SVEs and SDKIEs for WT and variant ssIGPS enzymes at 75 °C indicates that single amino acid substitutions can change the rate-determining step and hence what kcat reports on. A change in the rate-determining step also means that changes to kcat may underreport the impact of amino acid substitutions on the individual kinetic steps.