Synthesis and Degradation of Carbonic Anhydrase in a Synchronized Culture of Chlamydomonas reinhardtii

Abstract

The biosynthesis and degradation of carbonic anhydrase (CA; EC 4.3.1.1) was investigated during the course of synchronous culture of the unicellular green alga Chlamydomonas reinhardtii, carried out under a regime of 12 h of light and 12 h of darkness with bubbling of ordinary air. The enzymatic activity increased linearly during the light phase. A coordinate increase in the level of the 35-kDa mature CA polypeptide was demonstrated by immunostaining after poly-acrylamide gel electrophoresis and Western blotting. Pulse-labeling with [14C]arginine followed by immunoprecipitation showed that the biosynthesis of the CA polypeptide is very active in the early light phase and rapidly decreases after the middle of the light phase, indicating that the bio-synthetic activity does not reflect the quantity of enzyme protein or the level of enzymatic activity. The 42-kDa precursor but not the 35-kDa mature polypeptide was synthesized in the dark. The 35-kDa polypeptide, pulse-labeled at the beginning of the light phase, was gradually degraded throughout the light phase while it appeard to be stable in the dark. These results suggest that the messenger RNA coding for CA is present but its translation is limited in the dark. Normal translation of mRNA and processing of the precursor to yield the holoenzyme may both require light.