Biosynthesis and assembly of the proton-translocating adenosine triphosphatase complex from chloroplasts

Abstract

The H+-translocating ATPase complex of chloroplasts consists of at least 8 nonidentical subunits. Five of these (.alpha., .beta., .gamma., .delta., and .epsilon. subunits) collectively constitute the globular extramembranous CF1 (chloroplast coupling factor 1) portion of the complex. The remaining three subunits (I-III) represent the membrane-embedded portion. Biosynthesis and assembly of these subunits were studied by pulse-labeling isolated spinach chloroplasts in the presence of cycloheximide or chloramphenicol and by translating total leaf RNA in a rabbit reticulocyte system. The labeled products were analyzed by immunoprecipitation with subunit-specific antisera or by isolating the entire H+-translocating ATPase complex in a nearly pure state. Chloroplasts synthesize the .alpha., .beta., .gamma., and .epsilon. subunits of CF1, the membrane-embedded subunit I, and probably also the membrane-embedded subunit III. The .delta. subunit (and probably also subunit II) are imported from the cytoplasm via larger precursor forms. After isolated chloroplasts are labeled in the presence of cycloheximide, the chloroplast-made H+-ATPase subunits are assembled into a complex that is indistinguishable from the authentic H+-ATPase complex. Apparently isolated chloroplasts contain excess pools of the cytoplasmically made subunits.