Simultaneous determination of Rubisco carboxylase and oxygenase kinetic parameters in Triticum aestivum and Zea mays using membrane inlet mass spectrometry

The lack of complete Rubisco kinetic data for numerous species is partly because of the time consuming nature of the multiple methods needed to assay all of the Rubisco parameters. We have developed a membrane inlet mass spectrometer method that simultaneously determines the rate of Rubisco carboxylation (vc) and oxygenation (vo), and the CO2 and O2 concentrations. Using the collected data, the Michaels-Menten equations for vc and vo in response to changing CO2 and O2 concentrations were simultaneously solved for the CO2 (Kc) and O2 (Ko) constants, the maximum turnover rates of the enzyme for CO2 (kcatCO2) and O2 (kcatO2) and the specificity for CO2 relative to O2 (Sc/o). In the C4 species Zea mays Kc was higher but Ko was lower compared with the C3 species Triticum aestivum. The kcatCO2 was higher and the kcatO2 lower in Z. mays compared with T. aestivum and Sc/o was similar in the two species. The Vomax/Vcmax was lower in Z. mays and thus did not correlate with changes in Sc/o. In conclusion, this mass spectrometer system provides a means of simultaneously determining the important Rubisco kinetic parameters, Kc, Ko, kcatCO2,kcatO2 and Sc/o from the same set of assays.