The dynamics of SecM-induced translational stalling.
SecM is an E. coli secretion monitor capable of stalling translation on the prokaryotic ribosome without cofactors. Biochemical and structural studies have demonstrated that the SecM nascent chain interacts with the 50S subunit exit tunnel to inhibit peptide bond formation. However, the timescales and pathways of stalling on an mRNA remain undefined. To provide a dynamic mechanism for stalling, we directly tracked the dynamics of elongation on ribosomes translating the SecM stall sequence (FSTPVWISQAQGIRAGP) using single-molecule fluorescence techniques. Within 1 min, three peptide-ribosome interactions work cooperatively over the last five codons of the SecM sequence, leading to severely impaired elongation rates beginning from the terminal proline and lasting four codons. Our results suggest that stalling is tightly linked to the dynamics of elongation and underscore the roles that the exit tunnel and nascent chain play in controlling fundamental steps in translation.
Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-5126, USA; Department of Applied Physics, Stanford University, Stanford, CA 94305-4090, USA.
Albert
A
Tsai
Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-5126, USA.
Guy
G
Kornberg
Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-5126, USA.
Magnus
M
Johansson
Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-5126, USA; Department of Applied Physics, Stanford University, Stanford, CA 94305-4090, USA.
Jin
J
Chen
Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-5126, USA; Stanford Magnetic Resonance Laboratory, Stanford University School of Medicine, Stanford, CA 94305-5126, USA. Electronic address: puglisi@stanford.edu.
Joseph D
JD
Puglisi
D000595:Amino Acid Sequence; D003062:Codon / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010441:Peptide Chain Elongation, Translational; D012270:Ribosomes / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D003062:Codon; D029968:Escherichia coli Proteins; C497347:SecM protein, E coli; D014157:Transcription Factors