The amino-terminal 118 amino acids of Escherichia coli trigger factor constitute a domain that is necessary and sufficient for binding to ribosomes.
Escherichia coli trigger factor has prolyl-isomerase and chaperone activities and associates with nascent polypeptide chains. Trigger factor has a binding site on ribosomes, which is a prerequisite for its efficient association with nascent chains and its proposed function as a cotranslational folding catalyst. We set out to identify the domain of trigger factor that mediates ribosome binding. Of a series of recombinant fragments, the amino-terminal fragments, TF (1-144) and TF (1-247), cofractionated with ribosomes from cell extracts and rebound to isolated ribosomes in vitro. They competed efficiently with full-length trigger factor for stoichiometric binding to a single site on the large ribosomal subunit. However, TF (1-144) and TF (1-247) differed from full-length trigger factor in that their association with ribosomes was not strengthened by the presence of nascent chains, indicating a role for carboxyl-terminal trigger factor segment in sensing the translational status. The domain responsible for ribosome binding was further investigated by limited proteolysis of recombinant fragments. A stable domain comprising the amino-terminal 118 residues was identified that was still capable of ribosome binding and thus represents a novel structural and functional element of trigger factor.
Hesterkamp
T
T
Zentrum für Molekulare Biologie, Universität Heidelberg, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany.
Deuerling
E
E
Bukau
B
B
D000591:Amino Acid Isomerases / Q000737:chemistry / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology*; D010446:Peptide Fragments / Q000302:isolation & purification; D010449:Peptide Mapping; D019696:Peptidylprolyl Isomerase; D014176:Protein Biosynthesis; D017510:Protein Folding; D011691:Puromycin / Q000494:pharmacology; D012270:Ribosomes / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology
D001426:Bacterial Proteins; D002352:Carrier Proteins; D010446:Peptide Fragments; D012965:Sodium Chloride; D011691:Puromycin; D004706:Endodeoxyribonucleases; D000591:Amino Acid Isomerases; D019696:Peptidylprolyl Isomerase