Global profiling of SRP interaction with nascent polypeptides

• Published in: Nature (London) Vol. 536; no. 7615; pp. 219 - 223
• Main Authors: Schibich, Daniela, Gloge, Felix, Pöhner, Ina, Björkholm, Patrik, Wade, Rebecca C., von Heijne, Gunnar, Bukau, Bernd, Kramer, Günter
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 11-08-2016; Nature Publishing Group
• Subjects: 631/1647/514/2254; 631/337/574/1789; 631/45/612/1981; Amino acids; Binding Sites; Datasets; E coli; Escherichia coli - genetics; Escherichia coli - metabolism; Escherichia coli Proteins - biosynthesis; Escherichia coli Proteins - metabolism; Humanities and Social Sciences; Hydrophobic and Hydrophilic Interactions; letter; Localization; Membrane proteins; Membrane Proteins - biosynthesis; Membrane Proteins - metabolism; Membranes; multidisciplinary; Peptides - metabolism; Peptidylprolyl Isomerase - metabolism; Periplasm - metabolism; Physiological aspects; Polypeptides; Protein Binding; Protein Biosynthesis; Protein Structure, Tertiary; Protein Transport; Proteins; Proteome - biosynthesis; Proteome - metabolism; Proteomics; Ribosomes - metabolism; RNA; RNA, Bacterial - metabolism; RNA, Messenger - metabolism; Science; Signal Recognition Particle - metabolism; Substrate Specificity; Substrates; Translocation
• ISSN: 0028-0836; 1476-4687; 1476-4687
• DOI: 10.1038/nature19070

Here, the selection of substrates by the protein–RNA complex known as the signal recognition particle (SRP) is investigated in the bacterium
Escherichia coli
, revealing that the SRP has a strong preference for hydrophobic transmembrane domains of inner membrane proteins.
Specificity of the signal recognition particle
As nascent proteins are generated by translating ribosomes, they are simultaneously targeted for translocation into the endoplasmic reticulum by a protein–RNA complex known as the signal recognition particle (SRP). Günter Kramer and colleagues investigate the nature of SRP substrates and how are they selected in a study of the SRP interactome in the bacterium
Escherichia coli
. They find that SRP almost exclusively targets hydrophobic transmembrane domains (TMDs) of inner membrane proteins, rejecting proteins of the outer membrane and the periplasmic space. Their data also contradict previous thinking that SRP associates with the first TMD to emerge from a ribosome, and show that in many proteins that will eventually span the membrane several times, the SRP binds several TMDs. In a related paper also in this issue of
Nature
, Judith Frydman and co-workers, studying yeast cells, find that the SRP preferentially binds substrates destined for secretion before they are fully translated — specifically, through the non-coding elements in their mRNA and before 'targeting signals' of these substrates are translated.
Signal recognition particle (SRP) is a universally conserved protein–RNA complex that mediates co-translational protein translocation and membrane insertion by targeting translating ribosomes to membrane translocons
1
. The existence of parallel co- and post-translational transport pathways
2
, however, raises the question of the cellular substrate pool of SRP and the molecular basis of substrate selection. Here we determine the binding sites of bacterial SRP within the nascent proteome of
Escherichia coli
at amino acid resolution, by sequencing messenger RNA footprints of ribosome–nascent-chain complexes associated with SRP. SRP, on the basis of its strong preference for hydrophobic transmembrane domains (TMDs), constitutes a compartment-specific targeting factor for nascent inner membrane proteins (IMPs) that efficiently excludes signal-sequence-containing precursors of periplasmic and outer membrane proteins. SRP associates with hydrophobic TMDs enriched in consecutive stretches of hydrophobic and bulky aromatic amino acids immediately on their emergence from the ribosomal exit tunnel. By contrast with current models, N-terminal TMDs are frequently skipped and TMDs internal to the polypeptide sequence are selectively recognized. Furthermore, SRP binds several TMDs in many multi-spanning membrane proteins, suggesting cycles of SRP-mediated membrane targeting. SRP-mediated targeting is not accompanied by a transient slowdown of translation and is not influenced by the ribosome-associated chaperone trigger factor (TF), which has a distinct substrate pool and acts at different stages during translation. Overall, our proteome-wide data set of SRP-binding sites reveals the underlying principles of pathway decisions for nascent chains in bacteria, with SRP acting as the dominant triaging factor, sufficient to separate IMPs from substrates of the SecA–SecB post-translational translocation and TF-assisted cytosolic protein folding pathways.