title: Chain dynamics of nascent polypeptides emerging from the ribosome.
abstract: Very little is known about the conformation of polypeptides emerging from the ribosome during protein biosynthesis. Here, we explore the dynamics of ribosome-bound nascent polypeptides and proteins in Escherichia coli by dynamic fluorescence depolarization and assess the population of cotranslationally active chaperones trigger factor (TF) and DnaK. E. coli cell-free technology and fluorophore-linked E. coli Met-tRNA f Met enable selective site-specific labeling of nascent proteins at the N-terminal methionine. For the first time, direct spectroscopic evidence captures the generation of independent nascent chain motions for a single-domain protein emerging from the ribosome (apparent rotational correlation time approximately 5 ns), during the intermediate and late stages of polypeptide elongation. Such motions are detected only for a sequence encoding a globular protein and not for a natively unfolded control, suggesting that the independent nascent chain dynamics may be a signature of folding-competent sequences. In summary, we observe multicomponent, severely rotationally restricted, and strongly chain length/sequence-dependent nascent chain dynamics.
authors: Department of Chemistry, University of Wisconsin-Madison, 1101 University Avenue, Madison, Wisconsin 53706, USA.
authors: Jamie P
authors: JP
authors: Ellis
authors: Courtney K
authors: CK
authors: Bakke
authors: Robert N
authors: RN
authors: Kirchdoerfer
authors: Lisa M
authors: LM
authors: Jungbauer
authors: Silvia
authors: S
authors: Cavagnero
mesh_terms: D001059:Apoproteins / Q000096:biosynthesis*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000502:physiology; D005454:Fluorescence Polarization; D009211:Myoglobin / Q000096:biosynthesis*; D010455:Peptides / Q000378:metabolism*; D019696:Peptidylprolyl Isomerase / Q000502:physiology; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012270:Ribosomes / Q000378:metabolism*
chemical_list: D001059:Apoproteins; D029968:Escherichia coli Proteins; D009211:Myoglobin; D010455:Peptides; C010857:apomyoglobin; C403324:trigger factor, E coli; D019696:Peptidylprolyl Isomerase