The relationship between the three dimensional structure and the function of an enzyme is still poorly understood. This is especially so in the context of predictably altering the function of an enzyme. The interactive computer graphics in the Computer Graphics Laboratory provide a resource with which the binding surfaces and catalytic active sites can be discovered and mapped. This project involves creating and analyzing an altered serine protease. Granzyme B is a serine protease that had been implicated in cytotoxic lymphocyte mediated cell death. Experimenta evidence shows that granzyme B had strict extended specificity with the unusual property of cleaving after an acidic residue. The well characterized protease trypsin will be used as a model system to explore the specificity of granzyme B and other proteases with strict extended sites. Through three dimensional and sequence comparison the chymotrypsin family of enzymes will be analyzed for common motifs in extended specificity. When common themes occur, they will be tested on a hybrid scaffold for changes in Km. A hybrid will be designed that takes into account differences in the granzyme B and trypsin active sites. Trypsin has both a characteristic loop region and a disulfide bridge that is absent in granzyme B and may account for the differences in specificity. Mutations will be made systematically to trypsin to convert it to a protease of granzyme B specificity. Molecular modeling with Midas Plus and computer visualization will play important roles in the experimental design towards understanding the differences between these two and other serine proteases.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001081-21
Application #
6280296
Study Section
Project Start
1998-07-01
Project End
1999-06-30
Budget Start
1997-10-01
Budget End
1998-09-30
Support Year
21
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
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