IDENTIFICATION OF DISTINCT INTERNAL AND EXTERNAL ISOZYMES OF CARBONIC-ANHYDRASE IN CHLORELLA-SACCHAROPHILA

External carbonic anhydrase (CA) was detected in whole cells of alkali ne-grown Chlorella saccharophila but was suppressed by growth at acid pH or growth on elevated levels of CO2. Internal CA activity was measu red potentiometrically as an increase in activity in cell extracts ove r that of intact cells. Cells grown under all conditions had equal lev els of internal CA activity. Two isozymes were identified after electr ophoretic separation of soluble proteins on cellulose acetate plates. The fast isozyme was found in cells grown under all conditions, wherea s the slow isozyme was found only in cells grown at alkaline pH. Weste rn blot analysis following sodium dodecyl sulfate-polyacrylamide gel e lectrophoresis using antibodies produced against the periplasmic form of CA from Chlamydomonas reinhardtii revealed a single band at 39 kD, which did not change in intensity between growth conditions and was as sociated only with proteins eluted from the fast band. The slow isozym e was inactivated by incubation of cell extract at 30 degrees C and by incubation in 10 mM dithiothreitol, whereas the internal form was una ffected. These results indicate that external and internal forms of CA differ in structure and their activities respond differently to envir onmental conditions.