Signal Recognition Particle-ribosome Binding Is Sensitive to Nascent Chain Length

Noriega, Thomas R.; Tsai, Albert; Elvekrog, Margaret; Petrov, Alexey; Neher, Saskia B.; Chen, Jin; Bradshaw, Niels; Puglisi, Joseph D.; Walter, Peter

doi:10.1074/jbc.m114.563239

Cited by 41 publications

(51 citation statements)

References 44 publications

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“…Indeed, slower translation elongation, which gives SRP a longer time window, allows substrates with weaker signal sequences to enter the SRP pathway (75). The results of this and previous work (47,57,58) show that multiple mechanisms together impose this timer. First, RNC-SRP binding affinity could weaken modestly with long-chain RNCs.…”

Section: Discussionsupporting

confidence: 55%

“…Our findings also suggest a remarkable degree of structural plasticity for cotranslational biogenesis machineries bound at the ribosome exit site, wherein each factor could alter its conformation to accommodate another factor bound in the vicinity. In support of this model, multiple conformations of SRP bound to RNCs have been observed recently in fluorescence and single-molecule studies (34,58). Regulation of SRP conformation by TF presumably leads to the observed weaker binding to RNCs and the slower recruitment of FtsY in the membrane-targeting step.…”

Section: Discussionmentioning

confidence: 56%

“…S4). Importantly, the SRP-RNC 130-135 binding affinities observed here (Table 1) and elsewhere (58) are still in the low nanomolar range and sufficient for SRP to occupy the RNC at its cellular concentration. Likewise, a longer nascent chain slows SRP•FtsY closed complex assembly by less than threefold (Table 2).…”

Section: Resultsmentioning

confidence: 88%

“…However, the molecular basis underlying this phenomenon has been controversial. Some studies suggest that RNC-SRP binding affinity is reduced as the nascent chain becomes longer, whereas others suggest that SRP loses the ability to target longer nascent chains to the membrane (47,57,58).…”

Section: Resultsmentioning

confidence: 99%

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Regulation by a chaperone improves substrate selectivity during cotranslational protein targeting

Ariosa

Lee

Wang

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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The ribosome exit site is a crowded environment where numerous factors contact nascent polypeptides to influence their folding, localization, and quality control. Timely and accurate selection of nascent polypeptides into the correct pathway is essential for proper protein biogenesis. To understand how this is accomplished, we probe the mechanism by which nascent polypeptides are accurately sorted between the major cotranslational chaperone trigger factor (TF) and the essential cotranslational targeting machinery, signal recognition particle (SRP). We show that TF regulates SRP function at three distinct stages, including binding of the translating ribosome, membrane targeting via recruitment of the SRP receptor, and rejection of ribosome-bound nascent polypeptides beyond a critical length. Together, these mechanisms enhance the specificity of substrate selection into both pathways. Our results reveal a multilayered mechanism of molecular interplay at the ribosome exit site, and provide a conceptual framework to understand how proteins are selected among distinct biogenesis machineries in this crowded environment.signal recognition particle | trigger factor | ribosome | protein biogenesis | GTPases P roper protein biogenesis is a prerequisite for the maintenance of a functional proteome. Accumulating data indicate that this process begins at the ribosome exit site, where many protein biogenesis machineries can interact and gain access to the nascent polypeptide. This includes chaperones (1-5) such as trigger factor (TF) (1, 4, 6, 7), Hsp70, and the nascent polypeptide-associated complex (8-13); modification enzymes (10, 14-16) such as N-acetyl transferase, methionine aminopeptidase, and arginyl transferase; protein-targeting and translocation machineries such as signal recognition particle (SRP) (17-20), SecA (21), the SecYEG (or Sec61p) (22, 23) and YidC translocases (24,25), and the ribosome-bound quality control complex (26)(27)(28)(29)(30). Engagement of these factors with nascent polypeptides influences their folding, assembly, localization, processing, and quality control. Within seconds after the nascent polypeptide emerges from the ribosomal exit tunnel, it must engage the correct set of factors and thus commit to the proper biogenesis pathway. How this is accomplished in the crowded environment at the ribosome exit site is an emerging question. In this work, we address this question by deciphering how nascent proteins are selected between two major protein biogenesis machineries in bacteria, SRP and TF.SRP is a universally conserved ribonucleoprotein complex responsible for the cotranslational targeting of proteins to the eukaryotic endoplasmic reticulum (ER), or the bacterial plasma membrane (31). SRP recognizes ribosome-nascent chain complexes (termed RNC or cargo) carrying strong signal sequences and delivers them to the SecYEG or YidC translocation machinery on the target membrane. SRP binds RNC via two interactions: a helical N domain in the SRP54 protein (called Ffh in bacteria) binds the ribosoma...

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Section: Discussionsupporting

confidence: 55%

Section: Discussionmentioning

confidence: 56%

Section: Resultsmentioning

confidence: 88%

Section: Resultsmentioning

confidence: 99%

See 2 more Smart Citations

Regulation by a chaperone improves substrate selectivity during cotranslational protein targeting

Ariosa

Lee

Wang

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…It is not straightforward to draw conclusions about the M-domain conformation from these structures, because RNCs with a different nascent chain lengths and different signal sequences have been used in each study. Different signal sequence composition and nascent chain length has been reported to affect SRP binding to the ribosome (36). Therefore, it could be that the relatively short Lep50 nascent chain used in this study induces a less-flexible M domain, which thus can be observed in the EM density.…”

Section: Discussionmentioning

confidence: 92%

Ribosome–SRP–FtsY cotranslational targeting complex in the closed state

Loeffelholz

Jiang

Ariosa

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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The signal recognition particle (SRP)-dependent pathway is essential for correct targeting of proteins to the membrane and subsequent insertion in the membrane or secretion. In Escherichia coli, the SRP and its receptor FtsY bind to ribosome-nascent chain complexes with signal sequences and undergo a series of distinct conformational changes, which ensures accurate timing and fidelity of protein targeting. Initial recruitment of the SRP receptor FtsY to the SRP-RNC complex results in GTP-independent binding of the SRP-FtsY GTPases at the SRP RNA tetraloop. In the presence of GTP, a closed state is adopted by the SRP-FtsY complex. The cryo-EM structure of the closed state reveals an ordered SRP RNA and SRP M domain with a signal sequence-bound. Van der Waals interactions between the finger loop and ribosomal protein L24 lead to a constricted signal sequence-binding pocket possibly preventing premature release of the signal sequence. Conserved M-domain residues contact ribosomal RNA helices 24 and 59. The SRP-FtsY GTPases are detached from the RNA tetraloop and flexible, thus liberating the ribosomal exit site for binding of the translocation machinery.protein targeting | signal recognition particle | signal sequence | ribosome | single-particle electron cryomicroscopy T he Escherichia coli signal recognition particle (SRP) is a complex consisting of the universally conserved protein Ffh and 4.5S RNA, which adopts a hairpin structure (1). Ffh is composed of the N-terminal domain, the G domain that harbors GTPase activity, and the C-terminal methionine-rich M domain that interacts with 4.5S RNA (2, 3) and with the signal sequence (4, 5). The N and G domains form a compact structural and functional unit termed "the NG domain." Targeting of ribosome-nascent chain complexes (RNC) containing a signal sequence depends on the interaction of the RNC-SRP complex with the SRP receptor FtsY, which is membrane associated (6-9). FtsY and Ffh interact via their homologous NG domains and form a composite GTPase active site (10, 11). Crystal structures of the M domain reveal a hydrophobic groove used to capture signal sequences (4,5,12).Protein targeting is driven by highly regulated conformational rearrangements of SRP and FtsY as well as GTP hydrolysis. SRP recognizes and tightly binds to RNCs displaying a signal sequence (cargo). Next, RNC-bound SRP efficiently recruits FtsY to form a nucleotide-independent, transient early state that rearranges to a GTP-stabilized closed state (13). Ultimately, in the activated state, handover of the RNC to the Sec translocon takes place, followed by GTP hydrolysis and disassembly of the SRP-FtsY complex (14-16). These distinct conformational transitions are regulated by the ribosome and translocon in the membrane, leading to a switch from cargo recognition by SRP to cargo release (17, 18).Cryo-EM structures of bacterial SRP-bound RNCs revealed a tight cargo-recognition complex (19,20). In the SRP-FtsY early complex an overall detachment of SRP from the ribosome was observed (21). In this sta...

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The Archaeal Signal Recognition Particle: Present Understanding and Future Perspective

2016

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The signal recognition particle (SRP) and its receptor constitute universally conserved and essential cellular machinery that controls the proper membrane localization of nascent polypeptides with the transmembrane domain. In the past decade, there has been an immense advancement in our understanding of this targeting machine in all three domains of life. A significant portion of such progress came from the structural analysis of archaeal SRP components. Despite the availability of structural insights from different archaeal SRP components, little is known about protein translocation in this domain of life compared to either bacteria or eukaryotes. One of the primary reasons being limited availability of the genetic and cell biological tools in archaea. In the present review, an attempt has been made to explore the structural information available for archaeal SRP components to gain insights into the protein translocation mechanism of this group of organisms. Besides, many exciting avenues of archaeal research possible using the recently developed genetic and cell biological tools for some species have been identified.

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Signal Recognition Particle-ribosome Binding Is Sensitive to Nascent Chain Length

Cited by 41 publications

References 44 publications

Regulation by a chaperone improves substrate selectivity during cotranslational protein targeting

Regulation by a chaperone improves substrate selectivity during cotranslational protein targeting

Ribosome–SRP–FtsY cotranslational targeting complex in the closed state

The Archaeal Signal Recognition Particle: Present Understanding and Future Perspective

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