Our long term goal is to develop and exploit new strategies for the design and synthesis of biological catalysts with tailored specificities.
The specific aim of this proposal is to design, synthesize, and characterize sequence-specific hybrid ribonucleases and deoxyribonucleases with defined binding site sequences and binding site sizes. A new binding site consisting of an oligodeoxyribonucleotide of defined length and sequence has been site-selectively introduced outside the catalytic sites of the relatively nonspecific enzymes, bovine pancreatic ribonuclease A and staphylococcal nuclease. The resulting hybrid enzymes are able to sequence-specifically cleave single-stranded DNA and RNA. The design of this first generation of hybrid enzymes will be optimized and the catalytic properties and specificities characterized in detail. In addition, strategies will be explored for cleaving double- stranded DNA with these and other hybrid enzymes,including rec A mediated triple-strand formation, triple-helix formation, and derivatization of DNA binding proteins with staphylococcal nuclease. These hybrid sequence-specific phosphodiesterases will provide important tools for studies of both RNA and DNA structure and function as well as for mapping, isolating, and cloning nucleic acids. The strategies developed here may be applied to the design of other hybrid enzymes with tailored specificities for application in chemistry, biology, and medicine.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM041679-01
Application #
3300003
Study Section
Biochemistry Study Section (BIO)
Project Start
1989-04-01
Project End
1993-03-31
Budget Start
1989-04-01
Budget End
1990-03-31
Support Year
1
Fiscal Year
1989
Total Cost
Indirect Cost
Name
University of California Berkeley
Department
Type
Schools of Arts and Sciences
DUNS #
094878337
City
Berkeley
State
CA
Country
United States
Zip Code
94704
Sera, T; Schultz, P G (1996) In vivo selection of basic region-leucine zipper proteins with altered DNA-binding specificities. Proc Natl Acad Sci U S A 93:2920-5
Ku, J; Schultz, P G (1994) Phage display of catalytically active staphylococcal nuclease. Bioorg Med Chem 2:1413-5
Pei, D H; Ulrich, H D; Schultz, P G (1991) A combinatorial approach toward DNA recognition. Science 253:1408-11
Corey, D R; Pei, D; Schultz, P G (1989) Generation of a catalytic sequence-specific hybrid DNase. Biochemistry 28:8277-86