Synthesis of large subunit of ribulosebisphosphate carboxylase by thylakoid-bound polyribosomes from spinach chloroplasts.
Intact chloroplasts were isolated from developing first leaves of spinach. The chloroplasts were broken and separated into an extensively washed membrane (thylakoid) fraction and a soluble (stroma) fraction. The membrane fraction contained polyribosomes with properties similar to those of thylakoid-bound polyribosomes of other organisms. The distribution of mRNA for large-subunit ribulosebisphosphate carboxylase (LS) was determined by translating RNA from chloroplasts, thylakoids, and stroma in a wheat germ cell-free translation system. LS translation product was identified by immunoprecipitation with antibody to LS from spinach, electrophoresis of the immunoprecipitated product, and fluorography. At least 44% of translatable chloroplast LS-mRNA was in the washed thylakoid fraction. Thylakoid-bound LS-mRNA was in polyribosomes since LS was produced by thylakoids in an Escherichia coli cell-free translation system under conditions where initiation did not take place. Our results demonstrate that membrane-bound polyribosomes can synthesize the stroma-localized polypeptide LS, and suggest that the thylakoids may be an important site of its synthesis.
D011232:Chemical Precipitation; D002736:Chloroplasts / Q000201:enzymology*; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism*; D011132:Polyribosomes / Q000378:metabolism*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis; D012273:Ribulose-Bisphosphate Carboxylase / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D014357:Trypsin / Q000494:pharmacology; D014675:Vegetables
D008565:Membrane Proteins; D012333:RNA, Messenger; C021261:thylakoid polypeptides; D014357:Trypsin; D012273:Ribulose-Bisphosphate Carboxylase