Abstract
Within the large domain of the human signal recognition particle (SRP), 18 mutant SRP RNAs were constructed to disrupt Watson-Crick and G-U basepairs in helices 5, 6 and 8. Using a double-filter assay, the competitive binding of the mutant RNAs to purified human SRP68/72 or to a 7.4 kDa RNA-binding fragment of SRP72 (72frg) was measured. Binding of SRP68/72 was impaired by several mutations in the large domain with the most pronounced effects caused by changes in helix 5 (residues 222-231) and helix 8 (residues 176-191 and 202-214). Binding of the 72frg was diminished prominently by altering helix 5, in particular residues 120-128, and was unaffected by deleting helices 6 and 8. Deleting helix 8 diminished binding of SRP68/72 to a greater extent than deleting helix 6. The data suggest that nucleotide residues throughout most of the large SRP domain are directly and/or indirectly engaged in the binding of SRP68. In contrast, SRP72 binds only to a portion of the 5ef region.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 154-159 |
| Number of pages | 6 |
| Journal | RNA Biology |
| Volume | 4 |
| Issue number | 3 |
| State | Published - Jul 2007 |
| Externally published | Yes |
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Keywords
- Protein targeting
- Protein-RNA interactions
- RNP assembly
- Signal recognition particle
- Site-directed mutagenesis
- SRP
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology
Cite this
Identification of the RNA binding regions of SRP68/72 and SRP72 by systematic mutagenesis of human SRP RNA. / Yin, Jiaming; Iakhiaeva, Elena; Menichelli, Elena; Zwieb, Christian W.
In: RNA Biology, Vol. 4, No. 3, 07.2007, p. 154-159.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Identification of the RNA binding regions of SRP68/72 and SRP72 by systematic mutagenesis of human SRP RNA
AU - Yin, Jiaming
AU - Iakhiaeva, Elena
AU - Menichelli, Elena
AU - Zwieb, Christian W
PY - 2007/7
Y1 - 2007/7
N2 - Within the large domain of the human signal recognition particle (SRP), 18 mutant SRP RNAs were constructed to disrupt Watson-Crick and G-U basepairs in helices 5, 6 and 8. Using a double-filter assay, the competitive binding of the mutant RNAs to purified human SRP68/72 or to a 7.4 kDa RNA-binding fragment of SRP72 (72frg) was measured. Binding of SRP68/72 was impaired by several mutations in the large domain with the most pronounced effects caused by changes in helix 5 (residues 222-231) and helix 8 (residues 176-191 and 202-214). Binding of the 72frg was diminished prominently by altering helix 5, in particular residues 120-128, and was unaffected by deleting helices 6 and 8. Deleting helix 8 diminished binding of SRP68/72 to a greater extent than deleting helix 6. The data suggest that nucleotide residues throughout most of the large SRP domain are directly and/or indirectly engaged in the binding of SRP68. In contrast, SRP72 binds only to a portion of the 5ef region.
AB - Within the large domain of the human signal recognition particle (SRP), 18 mutant SRP RNAs were constructed to disrupt Watson-Crick and G-U basepairs in helices 5, 6 and 8. Using a double-filter assay, the competitive binding of the mutant RNAs to purified human SRP68/72 or to a 7.4 kDa RNA-binding fragment of SRP72 (72frg) was measured. Binding of SRP68/72 was impaired by several mutations in the large domain with the most pronounced effects caused by changes in helix 5 (residues 222-231) and helix 8 (residues 176-191 and 202-214). Binding of the 72frg was diminished prominently by altering helix 5, in particular residues 120-128, and was unaffected by deleting helices 6 and 8. Deleting helix 8 diminished binding of SRP68/72 to a greater extent than deleting helix 6. The data suggest that nucleotide residues throughout most of the large SRP domain are directly and/or indirectly engaged in the binding of SRP68. In contrast, SRP72 binds only to a portion of the 5ef region.
KW - Protein targeting
KW - Protein-RNA interactions
KW - RNP assembly
KW - Signal recognition particle
KW - Site-directed mutagenesis
KW - SRP
UR - http://www.scopus.com/inward/record.url?scp=39049124096&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=39049124096&partnerID=8YFLogxK
M3 - Article
C2 - 18347438
AN - SCOPUS:39049124096
VL - 4
SP - 154
EP - 159
JO - RNA Biology
JF - RNA Biology
SN - 1547-6286
IS - 3
ER -