Kinetic Characterization of Nitrite Uptake and Reduction by Chlamydomonas reinhardtii

Abstract

Kinetics of nitrite uptake and reduction by Chlamydomonas reinhardtii cells growing phototrophically has been studied by means of progress curves and the Michaelis-Menten integrated equation. Both uptake and reduction processes exhibited hyperbolic saturation kinetics, the nitrite uptake system lacking a diffusion component. Nitrite uptake and reduction showed significant differences in K_s for nitrite at pH 7.5 (1.6 versus 20 micromolar, respectively), optimal pH, activation energy values, and sensitivity toward reagents of sulfhydryl groups. K_s values for nitrite uptake were halved in cells subjected to darkness or to nitrogen-starvation. Nitrate inhibited nitrite uptake by a partially competitive mechanism. The same inhibition pattern was found for nitrite uptake by C. reinhardtii mutant 305 cells incapable of nitrate assimilation. The results demonstrate that C. reinhardtii cells take up nitrite via a highly specific carrier, probably energy-dependent, kinetically responsive to environmental changes, distinguishable from the enzymic nitrite reduction and endowed with an active site for nitrite not usable for nitrate transport.