Global Characterization of in Vivo Enzyme Catalytic Rates and Their Correspondence to in Vitro Kcat Measurements

Turnover Numbers, Also Known As Kcat Values, Are Fundamental Properties of Enzymes, However, Their Current Use Presents Two Major Difficulties. First, Many Values Were Never Experimentally Measured. Second, Most Experimentally-Available Kcat Values Were Measured Under Non-Physiological Conditions, That May Not Represent the in Vivo Situation. Here, We Estimate the Catalytic Rates Kapp of Enzymes in Vivo from Enzyme Abundance Data and Flux Predictions. By Taking the Maximum Across a Large Set of Cellular Conditions, We Calculate Kmax, the Observed Maximal Catalytic Rate of an Enzyme inside Cells, and Compare It to Kcat, the Maximal Rate of an Enzyme Measured in Laboratory Conditions. We Find a Log-Log Correlation of r²=0.61 (p<10^-10), with a Root Mean Square Difference of 0.55 (3.5-fold in linear scale), Indicating That Kcat Values and Maximal Rates in Vivo Generally Concur. to Investigate Differences Between in Vivo and in Vitro Rates, We Include Experimentally Measured Metabolite Concentrations to Calculate the Degree of Saturation of Enzymes and the Thermodynamic Driving Force. Correcting By These Effects Further Decreases the Typical Difference Between in Vivo and in Vitro Rates. the Presented Approach Offers a High Throughput Method for the Collection of Enzyme Kinetic Constants, and Unravels the Correspondence Between in Vitro and in Vivo Enzymatic Catalysis