Chloroplast envelope proteins from Chlamydomonas: separation into outer and inner envelopes and analysis by two-dimensional gel electrophoresis

Abstract

Fractions enriched in outer and inner chloroplast envelope membranes were isolated from the alga Chlamydomonas reinhardtii. Functionally active chloroplasts were lysed after equilibration in a hypertonic solution and outer and inner envelopes were separated. The protein content of each fraction was analysed by two-dimensional gel electrophoresis using isoelectric focusing/SDS polyacrylamide gel electrophoresis. The method is capable of separating components over a wide range of molecular mass and p I without interference by lipid binding. Each fraction contained a characteristic protein pattern. A 32 kDa protein was found exclusively associated with the inner envelope indicating that algal chloroplast envelopes share a common feature with those of higher plants.