New reagents for spectral titration of active site of chymotrypsin

H S Park; S Lee; J Suh

doi:10.1016/0165-022x(93)90007-b

New reagents for spectral titration of active site of chymotrypsin

J Biochem Biophys Methods. 1993 Oct;27(3):261-5. doi: 10.1016/0165-022x(93)90007-b.

Authors

H S Park¹, S Lee, J Suh

Affiliation

¹ Department of Chemistry, Seoul National University, South Korea.

PMID: 8258641
DOI: 10.1016/0165-022x(93)90007-b

Abstract

The reaction of alpha-chymotrypsin (ChT) with 2-phenyl-(E)-[4-(E)-cinnamylidene]oxazolin-5-one (E-PCO) or 2-phenyl-(Z)-[4-(E)-cinnamylidene]oxazolin-5-one (Z-PCO) at pH 4.5 and 25 degrees C led to the quantitative accumulation of the corresponding acyl-ChT intermediate. Very large changes in molar extinction coefficient were observed in the visible region during the conversion of the oxazolinones to the acyl-ChT intermediates. This afforded a reliable method for the spectral titration of ChT in the visible region. Both E-PCO and Z-PCO were stable under these conditions and did not cause any complication in the titration of the active site.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Chymotrypsin / analysis*
Indicators and Reagents / chemistry
Oxazoles / chemistry*
Spectrophotometry
Stereoisomerism

Substances

Indicators and Reagents
Oxazoles
2-phenyl-(4-cinnamylidene)oxazolin-5-one
Chymotrypsin