Kinetic measurement of serpin inhibitory activity by real-time fluorogenic biochemical assay

Jordan R. Yaron, Sriram Ambadapadi, Liqiang Zhang, Alexandra Lucas

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Scopus citations

Abstract

Biochemical fluorogenic and chromogenic assays facilitate real-time study of enzyme function. Based on the principle of enzymatic inhibition, these kinetic assays can be adapted to measure the function of serpins. Compared to traditional, electrophoretic study of serpin inhibitory complex formation, kinetic assays allow for finer temporal resolution as well as more quantitative comparisons between different conditions. This chapter describes methodology for performing real-time, kinetic measurement of serpin inhibitory activity by fluorogenic substrate conversion assay. Specifically, the methods covered include measurement of alpha-1-antitrypsin inhibitory activity against trypsin and heparin-dependent anti-thrombin III inhibitory activity against thrombin. These methods are scalable to small-volume, high-density format and can be applied for high-throughput screening of serpin activity modulators.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages65-71
Number of pages7
DOIs
StatePublished - 2018

Publication series

NameMethods in Molecular Biology
Volume1826
ISSN (Print)1064-3745

Keywords

  • Alpha-1-antitrypsin
  • Anti-thrombin III
  • Inhibition
  • Kinetic
  • Real-time
  • Serpin
  • Thrombin
  • Trypsin

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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