Membrane proteins synthesized but not processed by isolated maize chloroplasts

One-dimensional maps of proteolytic fragments generated by digestion with Staphylococcus aureus protease in sodium dodecyl sulfate (SDS) were used to identify three polypeptides synthesized by isolated Zea mays chloroplasts. This technique does not depend upon proper incorporation of the newly synthesized polypeptides into a more complex structure for their identification. The only preliminary purification required is electrophoretic separation on SDS-polyacrylamide gels. The pattern of radioactive fragments from labeled proteins which co-migrate with the alpha and beta subunits of chloroplast coupling factor (CF1) corresponds precisely to the pattern of stainable fragments derived from subunits of the purified enzyme. A 34,500-dalton protein is the major membrane-associated product of protein synthesis by isolated maize chloroplasts. From the similarity in the fragments formed by digestion with S. aureus protease, it appears that this radioactive protein is probably a precursor of a 32,000-dalton protein which is a component of the thylakoid. The alpha and beta subunits of CF1 newly synthesized by isolated chloroplasts are not fully extractable by procedures which normally solubilize the enzyme from membranes. The 34,500-dalton protein is not processed to the 32,000-dalton form in any great amount by isolated chloroplasts. A 19,000-dalton fragment of the 32,000-dalton protein is protected from digestion when thylakoids are treated with proteases, while the newly synthesized 34,500-dalton protein is fully susceptible. The isolated chloroplast does not appear to be able to fully integrate these newly made proteins into the membrane structure.

• Publication year

  1978

• Venue

  Journal of Cell Biology

• Publication date

  1978-09-01

• Fields of study

  Biology, Medicine, Environmental Science

• Identifiers
  DOI 10.1083/JCB.78.3.734PMID 701358PMCID 2110183
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• Newly synthesized CF1 alpha and beta subunits are not fully extractable by the usual membrane-solubilizing procedures, and the 34,500-dalton protein is not extensively processed to the 32,000-dalton form; isolated chloroplasts therefore do not fully integrate these products into the membrane.

  Confidence 0.93

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• The major membrane-associated translation product is a 34,500-dalton protein that is probably a precursor of the 32,000-dalton thylakoid protein.

  Confidence 0.95

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• Proteins co-migrating with the CF1 alpha and beta subunits yield radioactive fragment patterns that match the stainable fragments of the purified enzyme subunits exactly.

  Confidence 0.96

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• One-dimensional peptide mapping with Staphylococcus aureus protease can identify chloroplast-synthesized polypeptides from SDS-PAGE-separated proteins without requiring membrane integration into a larger complex.

  Confidence 0.94

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review

• 32,000-dalton protein

  protein

  A thylakoid-associated protein with an apparent mass of 32,000 daltons used for comparison in the fragment analysis.

  Aliases: 32 kDa protein

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• 34,500-dalton protein

  protein

  A membrane-associated chloroplast translation product with an apparent mass of 34,500 daltons.

  Aliases: 34.5 kDa protein

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• cf1 alpha subunit

  protein subunit

  The alpha polypeptide subunit of chloroplast coupling factor CF1.

  Aliases: alpha subunit of CF1

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• cf1 beta subunit

  protein subunit

  The beta polypeptide subunit of chloroplast coupling factor CF1.

  Aliases: beta subunit of CF1

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• isolated maize chloroplasts

  biological material

  Chloroplasts isolated from Zea mays and used as the protein-synthesizing system in the experiment.

  Aliases: isolated Zea mays chloroplasts, maize chloroplasts

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• membrane integration

  process

  The incorporation of newly synthesized proteins into chloroplast membrane structure.

  Aliases: integration into the membrane structure

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• one-dimensional peptide mapping

  method

  An analytical approach that compares proteolytic fragment patterns from proteins after SDS treatment.

  Aliases: one-dimensional maps of proteolytic fragments, peptide mapping

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• sds-page

  method

  Electrophoretic separation of proteins in SDS-polyacrylamide gels before fragment mapping.

  Aliases: sodium dodecyl sulfate-polyacrylamide gel electrophoresis

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• staphylococcus aureus protease

  enzyme

  A protease used here to digest SDS-denatured proteins into diagnostic fragments.

  Aliases: S. aureus protease

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review
• thylakoid

  membrane structure

  The internal chloroplast membrane system where the 32,000-dalton protein is a component.

  박진우 (dztg5apj7m) extraction뀨 (7c402c1b98) reviewAK (4715169a40) reviewB (s683577b42) reviewKiller Whale (322360f1c1) reviewq (76h6bfydm6) review

• Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis.

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  1973influential reference
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