Positive and negative mutant selection in the human histone hairpin-binding protein using the yeast three-hybrid system.

Martin, F; Michel, F; Zenklusen, D; Müller, B; Schümperli, Daniel (2000). Positive and negative mutant selection in the human histone hairpin-binding protein using the yeast three-hybrid system. Nucleic acids research, 28(7), pp. 1594-1603. Information Retrieval Ltd.

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We have used the yeast three-hybrid system in a positive selection for mutants of the human histone hairpin-binding protein (HBP) capable of interacting with non-canonical hairpins and in a negative selection for loss-of-binding mutants. Interestingly, all mutations from the positive selection are located in the N- and C-terminal regions flanking a minimal RNA-binding domain (RBD) previously defined between amino acids 126 and 198. Further, in vitro binding studies demonstrate that the RBD, which shows no obvious similarity to other RNA-binding motifs, has a relaxed sequence specificity compared to full-length HBP, allowing it to bind to mutant hairpin RNAs not normally found in histone genes. These findings indicate that the sequences flanking the RBD are important for restricting binding to the highly conserved histone hairpin structure. Among the loss-of-binding mutations, about half are nonsense mutations distributed throughout the N-terminal part and the RBD whereas the other half are missense mutations restricted to the RBD. Whereas the nonsense mutations permit a more precise definition of the C-terminal border of the RBD, the missense mutations identify critical residues for RNA binding within the RBD.

Item Type:

Journal Article (Original Article)

Division/Institute:

08 Faculty of Science > Department of Biology > Institute of Cell Biology
08 Faculty of Science > Department of Biology > Institute of Cell Biology > RNA

UniBE Contributor:

Schümperli, Daniel

Subjects:

500 Science > 570 Life sciences; biology
500 Science

ISSN:

0305-1048

Publisher:

Information Retrieval Ltd.

Language:

English

Submitter:

Daniel Schümperli

Date Deposited:

14 Sep 2015 10:08

Last Modified:

14 Sep 2015 10:08

PubMed ID:

10710426

URI:

https://boris.unibe.ch/id/eprint/71620

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