Hydrophobic core but not amino-terminal charged residues are required for translocation of an integral thylakoid membrane protein in vivo.
The integral membrane protein cytochrome f contains an amino-terminal signal sequence that is required for translocation into the thylakoid membrane. The signal sequence contains a hydrophobic core neighbored by an amino-terminal charged residue. Mutations that introduce charged amino acids into the hydrophobic core are inhibitory to cytochrome f translocation, and thus render cells non-photosynthetic. We have isolated both nuclear and chloroplast suppressors of these mutations by selecting for restoration of photosynthetic growth of Chlamydomonas. Here we describe the characterization of two chloroplast, second site suppressor mutations. Both suppressors remove the positively charged amino acid that borders the amino terminus of the hydrophobic core, and replace this arginine with either a cysteine or a leucine. The existence of these suppressors suggests that the hydrophobic core can be shifted in position within the signal sequence, and analysis of triple mutants in the signal confirms this hypothesis. Thus this signal that mediates translocation into the thylakoid membrane is characterized by a hydrophobic region whose exact amino acid content is not critical, and that need not be flanked on its amino terminus by a charged residue.
Baillet
B
B
Developmental Cell and Molecular Biology, Levine Science Research Center, Duke University, Durham, North Carolina 27708, USA.
Kohorn
B D
BD
D000595:Amino Acid Sequence; D000818:Animals; D001693:Biological Transport, Active; D002696:Chlamydomonas / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D003580:Cytochromes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D045348:Cytochromes f; D004563:Electrochemistry; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D021382:Protein Sorting Signals / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013489:Suppression, Genetic
D003580:Cytochromes; D008565:Membrane Proteins; D021382:Protein Sorting Signals; D045348:Cytochromes f