Structural analysis of cytochrome f from the psychrophillic alga Chlamydomonas subcaudata

Abstract

The psychrophillic green alga Chlamydomonas subcaudata was isolated from an Antarctic lake and appeared to be a natural state transition mutant. Cytochrome f (cyt f) from C. subcaudata has an apparent molecular mass which is 7kD lower than the apparent molecular mass of cyt f (41kD) from C. reinhardtii. Neither the cyt b6 nor the Rieske protein exhibit any differences in apparent molecular mass when both algal species were compared. We show that the apparent size difference between cyt f from C. subcaudata and C. reinhardtii is insensitive to the presence or absence of reducing reagents. However, the heme binding to cyt f from C. subcaudata is 2.3-fold less thermostable compared to that of C. reinhardtii. Analysis of cyt f from Arabidopsis (~32kD) and spinach (~40kD) showed that lower thermostability of heme binding to cyt f correlates with lower apparent molecular mass. Sequencing of the petA (cyt f coding region) from C. subcaudata revealed that it is identical in length and 77% identical in nucleotide sequence to the corresponding C. reinhardtii coding sequence. No substitutions of functionally important amino acids involved in the formation of the heme binding, water chain or plastocyanin interaction domains were found. Cyt f from both algal species were expressed in E. coli and the same difference in apparent molecular mass was observed. We suggest that the 7kD difference in the apparent molecular mass of cyt f from C. subcaudata compared to C. reinhardtii is due to a post-translational modification resulting in a conformational change.