Protein-induced Conformational Changes of RNA During the Assembly of Human Signal Recognition Particle

Elena, María; Isel, Catherine; Oubridge, Chris; Nagai, Kiyoshi

doi:10.1016/j.jmb.2006.12.056

Cited by 35 publications

(54 citation statements)

References 42 publications

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“…Thermodynamic cooperativity is important for the assembly of other multiprotein RNA complexes such as signal recognition particle or splicing complexes (Menichelli et al 2007;Wysoczanski et al 2014). The thermodynamic cooperativity between protein S4 and other ribosomal proteins measured in this study was comparable to that obtained in the previous studies on central domain proteins (Recht and Williamson 2001).…”

Section: Proteins S16 and S20 Destabilize The Intermediate S4-rrna Comentioning

confidence: 99%

Differential effects of ribosomal proteins and Mg²⁺ ions on a conformational switch during 30S ribosome 5′-domain assembly

Abeysirigunawardena

Woodson

2015

RNA

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Ribosomal protein S4 nucleates assembly of the 30S ribosome 5 ′ and central domains, which is crucial for the survival of cells. Protein S4 changes the structure of its 16S rRNA binding site, passing through a non-native intermediate complex before forming native S4-rRNA contacts. Ensemble FRET was used to measure the thermodynamic stability of non-native and native S4 complexes in the presence of Mg 2+ ions and other 5 ′ -domain proteins. Equilibrium titrations of Cy3-labeled 5 ′ -domain RNA with Cy5-labeled protein S4 showed that Mg 2+ ions preferentially stabilize the native S4-rRNA complex. In contrast, ribosomal proteins S20 and S16 act by destabilizing the non-native S4-rRNA complex. The full cooperative switch to the native complex requires S4, S16, and S20 and is achieved to a lesser degree by S4 and S16. The resulting thermodynamic model for assembly of the 30S body illustrates how ribosomal proteins selectively bias the equilibrium between alternative rRNA conformations, increasing the cooperativity of rRNA folding beyond what can be achieved by Mg 2+ ions alone.

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Section: Proteins S16 and S20 Destabilize The Intermediate S4-rrna Comentioning

confidence: 99%

Differential effects of ribosomal proteins and Mg²⁺ ions on a conformational switch during 30S ribosome 5′-domain assembly

Abeysirigunawardena

Woodson

2015

RNA

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“…We also investigated the full-length SRP68/72 complex, coexpressed in yeast and purified by Ni-NTA chromatography via the 6xHis tag attached to the Cterminus of SRP72 20 (''Materials and Methods'' section). In contrast to 68e 0 /H72b 0 , dissociation of 68/ 72H was observed over a wide range of urea concentrations.…”

Section: Disruption Of the Srp68/72 Interfacementioning

confidence: 99%

“…19 Consistent with these findings, chemical modification and footprinting experiments show that SRP68/72 brings helices 6 and 8 closer together to prepare the SRP RNA for the binding of the highly conserved SRP54 protein. 20 Because SRP72 binds only to a small section of helix 5, most RNA-protein contacts are likely to be brought about by SRP68. We have shown that a single conserved adenosine residue within the eukaryotic 5e motif is essential for the formation of a complex with human SRP72.…”

mentioning

confidence: 99%

Characterization of the SRP68/72 interface of human signal recognition particle by systematic site‐directed mutagenesis

Iakhiaeva

Hinck

et al. 2009

Protein Science

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The signal recognition particle (SRP) is a ribonucleoprotein complex which is crucial for the delivery of proteins to cellular membranes. Among the six proteins of the eukaryotic SRP, the two largest, SRP68 and SRP72, form a stable SRP68/72 heterodimer of unknown structure which is required for SRP function. Fragments 68e 0 (residues 530 to 620) and 72b 0 (residues 1 to 166) participate in the SRP68/72 interface. Both polypeptides were expressed in Escherichia coli and assembled into a complex which was stable at high ionic strength. Disruption of 68e 0 /72b 0 and SRP68/72 was achieved by denaturation using moderate concentrations of urea. The four predicted tetratricopeptide repeats (TPR1 to TPR4) of 72b 0 were required for stable binding of 68e 0 . Site-directed mutagenesis suggested that they provide the structural framework for the binding of SRP68. Deleting the region between TPR3 and TPR4 (h120) also prevented the formation of a heterodimer, but this predicted alpha-helical region appeared to engage several of its amino acid residues directly at the interface with 68e 0 . A 39-residue polypeptide (68h, residues 570-605), rich in prolines and containing an invariant aspartic residue at position 585, was found to be active. Mutagenesis scanning of the central region of 68h demonstrated that D585 was solely responsible for the formation of the heterodimer. Coexpression experiments suggested that 72b 0 protects 68h from proteolytic digestion consistent with the assertion that 68h is accommodated inside a groove formed by the superhelically arranged four TPRs of the N-terminal region of SRP72.

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“…[22][23][24][25][26] The assembly of the small bacterial SRPs, composed of an RNA (4.5S RNA) and one protein molecule (the SRP54 homologue Ffh), appears to be controlled by the expression levels and binding affinity between the two components. In eukaryotes, the SRP RNA was found to be located in a specialized region of the nucleolus where it combines with the imported SRP proteins SRP9/14, SRP19 and SRP68/72.…”

Section: 21mentioning

confidence: 99%

Kinship in the SRP RNA family

Rosenblad¹,

Larsen²,

Samuelsson³

et al. 2009

RNA Biology

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Protein-induced Conformational Changes of RNA During the Assembly of Human Signal Recognition Particle

Cited by 35 publications

References 42 publications

Differential effects of ribosomal proteins and Mg²⁺ ions on a conformational switch during 30S ribosome 5′-domain assembly

Differential effects of ribosomal proteins and Mg²⁺ ions on a conformational switch during 30S ribosome 5′-domain assembly

Characterization of the SRP68/72 interface of human signal recognition particle by systematic site‐directed mutagenesis

Kinship in the SRP RNA family

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Protein-induced Conformational Changes of RNA During the Assembly of Human Signal Recognition Particle

Cited by 35 publications

References 42 publications

Differential effects of ribosomal proteins and Mg2+ ions on a conformational switch during 30S ribosome 5′-domain assembly

Differential effects of ribosomal proteins and Mg2+ ions on a conformational switch during 30S ribosome 5′-domain assembly

Characterization of the SRP68/72 interface of human signal recognition particle by systematic site‐directed mutagenesis

Kinship in the SRP RNA family

Contact Info

Product

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Differential effects of ribosomal proteins and Mg²⁺ ions on a conformational switch during 30S ribosome 5′-domain assembly

Differential effects of ribosomal proteins and Mg²⁺ ions on a conformational switch during 30S ribosome 5′-domain assembly