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Biomedical subjects

Y Choo

Publications and source records attributed to Y Choo.

23 records · Page 2Linked to original sources
PubMed1994

In vivo repression by a site-specific DNA-binding protein designed against an oncogenic sequence.

A DNA-binding peptide comprising three zinc-fingers has been engineered to bind specifically to a unique nine-base-pair region of a BCR-ABL fusion oncogene in preference to the parent genomic sequences. Binding to the target oncogene in chromosomal DNA is possible in transformed cells in culture, and results in blockage of transcription. Consequently, murine cells rendered independent of growth factors by the action of the oncogene revert to factor dependence upon transient transfection with a vector expressing the peptide.

Amino Acid Sequence↗
PubMed1994

Toward a code for the interactions of zinc fingers with DNA: selection of randomized fingers displayed on phage.

We have used two selection techniques to study sequence-specific DNA recognition by the zinc finger, a small, modular DNA-binding minidomain. We have chosen zinc fingers because they bind as independent modules and so can be linked together in a peptide designed to bind a predetermined DNA site. In this paper, we describe how a library of zinc fingers displayed on the surface of bacteriophage enables selection of fingers capable of binding to given DNA triplets. The amino acid sequences of selected fingers which bind the same triplet are compared to examine how sequence-specific DNA recognition occurs. Our results can be rationalized in terms of coded interactions between zinc fingers and DNA, involving base contacts from a few alpha-helical positions. In the paper following this one, we describe a complementary technique which confirms the identity of amino acids capable of DNA sequence discrimination from these positions.

Amino Acid Sequence↗
PubMed1994

Selection of DNA binding sites for zinc fingers using rationally randomized DNA reveals coded interactions.

In the preceding paper [Choo, Y. & Klug, A. (1994) Proc. Natl. Acad. Sci. USA 91, 11163-11167], we showed how selections from a library of zinc fingers displayed on phage yielded fingers able to bind to a number of DNA triplets. Here, we describe a technique to deal efficiently with the converse problem--namely, the selection of a DNA binding site for a given zinc finger. This is done by screening against libraries of DNA triplet binding sites randomized in two positions but having one base fixed in the third position. The technique is applied here to determine the specificity of fingers previously selected by phage display. We find that some of these fingers are able to specify a unique base in each position of the cognate triplet. This is further illustrated by examples of fingers which can discriminate between closely related triplets as measured by their respective equilibrium dissociation constants. Comparing the amino acid sequences of fingers which specify a particular base in a triplet, we infer that in most instances, sequence-specific binding of zinc fingers to DNA can be achieved by using a small set of amino acid-nucleotide base contacts amenable to a code.

Amino Acid Sequence↗
PubMed1993

A role in DNA binding for the linker sequences of the first three zinc fingers of TFIIIA.

Zinc fingers of the TFIIIA type are connected by short linker sequences between the structural units. Structural investigations by 2D NMR in solution and by X-ray crystallographic analyses of complexes with DNA point to a passive role for the linkers. We have therefore investigated the influence of the linker sequence on DNA binding using as a model the first three fingers of the protein TFIIIA. Insertion of certain heterologous linkers abolishes binding, and replacement of individual amino acids can reduce binding by factors of up to twenty-four.

Amino Acid Sequence↗