[Frontiers in Bioscience 6, a13-16, April 1, 2001]

SINGLE MICROASSAY FOR MATRIX DEGRADING ENZYMES

Mahesh Mathrubutham and Srinivasa.K.Rao

Long Island Jewish Medical Center, Long Island campus for the Albert Einstein College of Medicine, 270-05 76th Avenue, New Hyde Park, New York 11042

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and Methods
3.1. Determination of Optimum Assay Buffer:
3.2. Effect of Optimum Buffer on Enzyme Rate of Reaction
3.3. Combination Substrate
4. Results
4.1. Optimum Assay Buffer
4.2. Effect of Combination Substrate
5. Discussion
6. References

1. ABSTRACT

Matrix degrading enzymes are implicated in several disease processes such as abdominal aortic aneurysms and emphysema, however, monitoring proteolytic activity in a single assay is not well-established. Numerous assays have been developed to measure matrix degrading enzymes, which use artificial substrates or substrates derived from natural substrate protein. We have recently developed an assay for elastolytic activity based on the detection of primary amines, using trinitrobenzene sulfonic acid (TNBSA), following the digestion of succinylated elastin. The assay is also versatile enough to allow the detection of other proteases through the use of succinylated substrate specific for given protease.