POLYPEPTIDE COMPOSITION, ASSEMBLY AND PHOSPHORYLATION PATTERNS OF THEPHOTOSYSTEM-II ANTENNA SYSTEM OF CHLAMYDOMONAS-REINHARDTII

In recent years major progress has been made in describing the gene fa milies that encode the polypeptides of the light-harvesting antenna sy stem of photosystem II (PSII). At the same time, advances in the bioch emical characterization of these antennae have been hampered by the hi gh degree of similarity between the apoproteins. To help interpret the molecular results, we have re-examined the composition, the assembly and the phosphorylation patterns of the light-harvesting antenna of PS II (LHCII) in the green alga Chlamydomonas reinhardtii Dang, using a n on-Tris SDS-PAGE system capable of resolving polypeptides that differ by as little as 200 daltons. Research to date has suggested that in C. reinhardtii the LHCII comprises just four polypeptides (p11, p13, p16 and p17), and CP29 and CP26 just one polypeptide each (p9 and p10, re spectively), i.e. a total of six polypeptides. We report here that the se antenna systems contain at least 15 polypeptides, 10 associated wit h LHCII, 3 with CP29, and 2 with CP26. All of these polypeptides have been positively identified by means of appropriate antibodies. We also demonstrate substantial heterogeneity to the pattern of in-vitro phos phorylation, with major differences found among members of closely spa ced and immunologically related polypeptides. Most intriguing is the f act that the polypeptides that cross-react with the anti-type 2 LHCII antibodies of higher plants (p16, and to a lesser extent p11) are not phosphorylated, whereas in higher plants these are the most highly pho sphorylated polypeptides. Also, unlike in higher plants, CP29 is heavi ly phosphorylated. Phosphorylation does not appear to have any effect on the mobility of polypeptides on fully denaturing SDS-PAGE gels. To learn more about the accumulation and organization of the light-harves ting polypeptides, we have also investigated a chlorophyll b-less muta nt, cbn1-48. The LHCII is almost completely lost in this mutant, along with at least some LHCI. But the accumulation of CP29 and CP26 and th eir binding to PSII core complexes, is relatively unaffected. As expec ted, the loss of antenna polypeptides is accompanied by a reduction of the size of large reaction-center complexes. Following in-vitro phosp horylation the number of phosphorylated proteins is greatly increased in the mutant thylakoids compared to wildtype thylakoids. We present a model of the PSII antenna system to account for the new polypeptide c omplexity we have demonstrated.