Towards a sequence-specific antibody protease

• Main Author: Smith, Robert Milton
• Language: ENG
• Published: ScholarWorks@UMass Amherst 1996
• Subjects: Biochemistry|Immunology
• Online Access: https://scholarworks.umass.edu/dissertations/AAI9709656

The exo-(1-amino-2-hydroxy)bicyclo (2.2,1) heptane-7-carboxylic acid derivatives I and II have been synthesized, the pivotal step being the use of an acyl nitrene-insertion reaction to introduce nitrogen functionality into the corresponding hydroxy ester. The analogues each mimic a distorted peptide ground state as well as the transition state for peptide bond hydrolysis. To enhance the immune response and to provide further sequence specificity, the analogues I and II have been coupled to a scD-Phe residue to yield the derivatives XIX, and XVII, respectively. Antibodies elicited against these derivatives may catalyze the hydrolysis of the corresponding peptides both by straining the substrate ground state and by stabilizing the transition state. A problem frequently associated with the detection of antibody catalysis of amide-bond hydrolysis is the difficulty of measuring small rate enhancements over the very slow rate of background hydrolysis. Thus, as a key component to this research, a method to accurately detect very low levels of free amines resulting from the background rate of peptide-bond hydrolysis has been advanced. This method utilizes fluorescence detection of naphthalenedicarboxaldehyde-derivatized peptide amines. As a test of this method, a pH-rate profile of the hydrolysis of N-(phenylacetyl)glycyl- scD-valine has been constructed.