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

2015 ◽  
Vol 112 (13) ◽  
pp. 3943-3948 ◽  
Author(s):  
Ottilie von Loeffelholz ◽  
Qiyang Jiang ◽  
Aileen Ariosa ◽  
Manikandan Karuppasamy ◽  
Karine Huard ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Protein Targeting ◽  
Signal Sequence ◽  
Signal Recognition Particle ◽  
Binding Pocket ◽  
Van Der Waals Interactions ◽  
Closed State ◽  
Chain Complexes ◽  
Nascent Chain ◽  
Srp Rna

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.

scholarly journals Structure, dynamics and interactions of large SRP variants

2019 ◽  
Vol 401 (1) ◽  
pp. 63-80 ◽  
Author(s):  
Klemens Wild ◽  
Matthias M.M. Becker ◽  
Georg Kempf ◽  
Irmgard Sinning
Keyword(s):  
Protein Targeting ◽  
Signal Recognition Particle ◽  
Domain Architecture ◽  
Particle Dynamics ◽  
Signal Recognition ◽  
Chain Complexes ◽  
Nascent Chain ◽  
Target Membrane ◽  
Signal Anchor ◽  
Srp Rna

Abstract Co-translational protein targeting to membranes relies on the signal recognition particle (SRP) system consisting of a cytosolic ribonucleoprotein complex and its membrane-associated receptor. SRP recognizes N-terminal cleavable signals or signal anchor sequences, retards translation, and delivers ribosome-nascent chain complexes (RNCs) to vacant translocation channels in the target membrane. While our mechanistic understanding is well advanced for the small bacterial systems it lags behind for the large bacterial, archaeal and eukaryotic SRP variants including an Alu and an S domain. Here we describe recent advances on structural and functional insights in domain architecture, particle dynamics and interplay with RNCs and translocon and GTP-dependent regulation of co-translational protein targeting stimulated by SRP RNA.

scholarly journals NAC functions as a modulator of SRP during the early steps of protein targeting to the endoplasmic reticulum

2012 ◽  
Vol 23 (16) ◽  
pp. 3027-3040 ◽  
Author(s):  
Ying Zhang ◽  
Uta Berndt ◽  
Hanna Gölz ◽  
Arlette Tais ◽  
Stefan Oellerer ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Targeting ◽  
Signal Sequence ◽  
Signal Recognition Particle ◽  
Signal Recognition ◽  
Signal Sequences ◽  
Chain Complexes ◽  
Ribosomal Tunnel ◽  
Nascent Chain ◽  
Combined Data

Nascent polypeptide-associated complex (NAC) was initially found to bind to any segment of the nascent chain except signal sequences. In this way, NAC is believed to prevent mistargeting due to binding of signal recognition particle (SRP) to signalless ribosome nascent chain complexes (RNCs). Here we revisit the interplay between NAC and SRP. NAC does not affect SRP function with respect to signalless RNCs; however, NAC does affect SRP function with respect to RNCs targeted to the endoplasmic reticulum (ER). First, early recruitment of SRP to RNCs containing a signal sequence within the ribosomal tunnel is NAC dependent. Second, NAC is able to directly and tightly bind to nascent signal sequences. Third, SRP initially displaces NAC from RNCs; however, when the signal sequence emerges further, trimeric NAC·RNC·SRP complexes form. Fourth, upon docking to the ER membrane NAC remains bound to RNCs, allowing NAC to shield cytosolically exposed nascent chain domains not only before but also during cotranslational translocation. The combined data indicate a functional interplay between NAC and SRP on ER-targeted RNCs, which is based on the ability of the two complexes to bind simultaneously to distinct segments of a single nascent chain.

scholarly journals The Signal Recognition Particle (SRP) RNA Links Conformational Changes in the SRP to Protein Targeting

2007 ◽  
Vol 18 (7) ◽  
pp. 2728-2734 ◽  
Author(s):  
Niels Bradshaw ◽  
Peter Walter
Keyword(s):  
Conformational Changes ◽  
Protein Targeting ◽  
Signal Sequence ◽  
Signal Recognition Particle ◽  
Signal Recognition ◽  
Gtp Hydrolysis ◽  
E Coli ◽  
Srp Rna ◽  
Binding Subunit

The RNA component of the signal recognition particle (SRP) is universally required for cotranslational protein targeting. Biochemical studies have shown that SRP RNA participates in the central step of protein targeting by catalyzing the interaction of the SRP with the SRP receptor (SR). SRP RNA also accelerates GTP hydrolysis in the SRP·SR complex once formed. Using a reverse-genetic and biochemical analysis, we identified mutations in the E. coli SRP protein, Ffh, that abrogate the activity of the SRP RNA and cause corresponding targeting defects in vivo. The mutations in Ffh that disrupt SRP RNA activity map to regions that undergo dramatic conformational changes during the targeting reaction, suggesting that the activity of the SRP RNA is linked to the major conformational changes in the signal sequence-binding subunit of the SRP. In this way, the SRP RNA may coordinate the interaction of the SRP and the SR with ribosome recruitment and transfer to the translocon, explaining why the SRP RNA is an indispensable component of the protein targeting machinery.

scholarly journals Efficient Interaction between Two GTPases Allows the Chloroplast SRP Pathway to Bypass the Requirement for an SRP RNA

2007 ◽  
Vol 18 (7) ◽  
pp. 2636-2645 ◽  
Author(s):  
Peera Jaru-Ampornpan ◽  
Sowmya Chandrasekar ◽  
Shu-ou Shan
Keyword(s):  
Complex Formation ◽  
Conformational Changes ◽  
Protein Targeting ◽  
Signal Recognition Particle ◽  
The Other ◽  
Signal Recognition ◽  
Binding Partner ◽  
Biochemical Analyses ◽  
Kinetics Of ◽  
Srp Rna

Cotranslational protein targeting to membranes is regulated by two GTPases in the signal recognition particle (SRP) and the SRP receptor; association between the two GTPases is slow and is accelerated 400-fold by the SRP RNA. Intriguingly, the otherwise universally conserved SRP RNA is missing in a novel chloroplast SRP pathway. We found that even in the absence of an SRP RNA, the chloroplast SRP and receptor GTPases can interact efficiently with one another; the kinetics of interaction between the chloroplast GTPases is 400-fold faster than their bacterial homologues, and matches the rate at which the bacterial SRP and receptor interact with the help of SRP RNA. Biochemical analyses further suggest that the chloroplast SRP receptor is pre-organized in a conformation that allows optimal interaction with its binding partner, so that conformational changes during complex formation are minimized. Our results highlight intriguing differences between the classical and chloroplast SRP and SRP receptor GTPases, and help explain how the chloroplast SRP pathway can mediate efficient targeting of proteins to the thylakoid membrane in the absence of the SRP RNA, which plays an indispensable role in all the other SRP pathways.

scholarly journals Protein Targeting to the Bacterial Cytoplasmic Membrane

1999 ◽  
Vol 63 (1) ◽  
pp. 161-173 ◽  
Author(s):  
Peter Fekkes ◽  
Arnold J. M. Driessen
Keyword(s):  
Protein Targeting ◽  
Cytoplasmic Membrane ◽  
Signal Sequence ◽  
Specific Interaction ◽  
Signal Recognition Particle ◽  
High Fidelity ◽  
Signal Recognition ◽  
Nascent Chain ◽  
Major Route ◽  
Mature Domain

SUMMARY Proteins that perform their activity within the cytoplasmic membrane or outside this cell boundary must be targeted to the translocation site prior to their insertion and/or translocation. In bacteria, several targeting routes are known; the SecB- and the signal recognition particle-dependent pathways are the best characterized. Recently, evidence for the existence of a third major route, the twin-Arg pathway, was gathered. Proteins that use either one of these three different pathways possess special features that enable their specific interaction with the components of the targeting routes. Such targeting information is often contained in an N-terminal extension, the signal sequence, but can also be found within the mature domain of the targeted protein. Once the nascent chain starts to emerge from the ribosome, competition for the protein between different targeting factors begins. After recognition and binding, the targeting factor delivers the protein to the translocation sites at the cytoplasmic membrane. Only by means of a specific interaction between the targeting component and its receptor is the cargo released for further processing and translocation. This mechanism ensures the high-fidelity targeting of premembrane and membrane proteins to the translocation site.

scholarly journals The many faces of SRP RNA

2014 ◽  
Vol 70 (a1) ◽  
pp. C1814-C1814
Author(s):  
Klemens Wild ◽  
Georg Kempf ◽  
Jan Grotwinkel ◽  
Irmgard Sinning
Keyword(s):  
Ternary Complex ◽  
Signal Recognition Particle ◽  
Signal Recognition ◽  
Chain Complexes ◽  
Composition And Structure ◽  
Nascent Chain ◽  
Domains Of Life ◽  
The Many ◽  
Srp Rna ◽  
High Diversity

The signal recognition particle (SRP) is a ribonucleoprotein complex that plays an essential role in co-translational targeting of membrane proteins. It is found in all three domains of life and exhibits a high diversity regarding composition and structure. In most organisms, SRP can be divided into two functional domains. The S domain mediates recognition and transport of ribosome-nascent chain complexes to the translocation channel, while the Alu domain stalls elongation of the ribosome until the complex has been faithfully delivered._x000B_Here we present the crystal structures of the complete bacterial SRP Alu domain and the ternary complex of human SRP S domain RNA, SRP19, and the SRP68-RBD. Together with previous structures, our data underline the taxon-specific evolutionary adaptation of SRP RNA that has important implications in SRP-mediated targeting.

SRP RNA Remodeling by SRP68 Explains Its Role in Protein Translocation

Science ◽  
2014 ◽  
Vol 344 (6179) ◽  
pp. 101-104 ◽  
Author(s):  
Jan Timo Grotwinkel ◽  
Klemens Wild ◽  
Bernd Segnitz ◽  
Irmgard Sinning
Keyword(s):  
Ribosomal Rna ◽  
Protein Targeting ◽  
Rna Binding ◽  
Protein Translocation ◽  
Signal Recognition Particle ◽  
Structural Basis ◽  
Signal Recognition ◽  
Rna Remodeling ◽  
Srp Rna ◽  
Arginine Rich Motif

The signal recognition particle (SRP) is central to membrane protein targeting; SRP RNA is essential for SRP assembly, elongation arrest, and activation of SRP guanosine triphosphatases. In eukaryotes, SRP function relies on the SRP68-SRP72 heterodimer. We present the crystal structures of the RNA-binding domain of SRP68 (SRP68-RBD) alone and in complex with SRP RNA and SRP19. SRP68-RBD is a tetratricopeptide-like module that binds to a RNA three-way junction, bends the RNA, and inserts an α-helical arginine-rich motif (ARM) into the major groove. The ARM opens the conserved 5f RNA loop, which in ribosome-bound SRP establishes a contact to ribosomal RNA. Our data provide the structural basis for eukaryote-specific, SRP68-driven RNA remodeling required for protein translocation.

scholarly journals Real-time observation of signal recognition particle binding to actively translating ribosomes

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Thomas R Noriega ◽  
Jin Chen ◽  
Peter Walter ◽  
Joseph D Puglisi
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Protein Translocation ◽  
Signal Sequence ◽  
Signal Recognition Particle ◽  
Initial Step ◽  
Signal Recognition ◽  
Fluorescence Measurements ◽  
Nascent Chain ◽  
Time Observation

The signal recognition particle (SRP) directs translating ribosome-nascent chain complexes (RNCs) that display a signal sequence to protein translocation channels in target membranes. All previous work on the initial step of the targeting reaction, when SRP binds to RNCs, used stalled and non-translating RNCs. This meant that an important dimension of the co-translational process remained unstudied. We apply single-molecule fluorescence measurements to observe directly and in real-time E. coli SRP binding to actively translating RNCs. We show at physiologically relevant SRP concentrations that SRP-RNC association and dissociation rates depend on nascent chain length and the exposure of a functional signal sequence outside the ribosome. Our results resolve a long-standing question: how can a limited, sub-stoichiometric pool of cellular SRP effectively distinguish RNCs displaying a signal sequence from those that are not? The answer is strikingly simple: as originally proposed, SRP only stably engages translating RNCs exposing a functional signal sequence.

scholarly journals Mechanisms of membrane protein biogenesis

2014 ◽  
Vol 70 (a1) ◽  
pp. C1161-C1161
Author(s):  
Irmgard Sinning
Keyword(s):  
Membrane Proteins ◽  
Signal Sequence ◽  
Signal Recognition Particle ◽  
Signal Recognition ◽  
Protein Biogenesis ◽  
Cargo Proteins ◽  
Target Membrane ◽  
Hydrophobic Nature ◽  
Correct Target ◽  
Srp Rna

More than 25% of the cellular proteome comprise membrane proteins that have to be inserted into the correct target membrane. Most membrane proteins are delivered to the membrane by the signal recognition particle (SRP) pathway which relies on the recognition of an N-terminal signal sequence. In contrast to this co-translational mechanism, which avoids problems due to the hydrophobic nature of the cargo proteins, tail-anchored (TA) membrane proteins utilize a post-translational mechanism for membrane insertion – the GET pathway (guided entry of tail-anchored membrane proteins). The SRP and GET pathways are both regulated by GTP and ATP binding proteins of the SIMIBI family. However, in the SRP pathway the SRP RNA plays a unique regulatory role. Recent insights into eukaryotic SRP will be discussed.

Sign in / Sign up

Export Citation Format

Share Document