Fluorescence spectroscopy analysis of active and regulatory sites of RNA polymerase

Abstract

This chapter discusses the fluorescence spectroscopy analysis of active and regulatory sites of RNA polymerase. The chapter divides the primary method of fluorescence spectroscopy into two categories to understand both the functional and the structural aspects of Escherichia coli (E. Coli) RNA polymerase. First, it deals with the extrinsic probe labeling of the substrate and enzyme, and then discusses the different important conclusions that can be drawn from such studies. Next, the time-resolved emission spectroscopic analysis of tryptophan fluorescence and the structure-function relationship of a transcription factor are discussed. The chapter discusses the syntheses and characterization of (γ-AmNS) ATP and UTP. The labeling of E. coli RNA polymerase and its subunits is discussed. The E. coli RNA polymerase has five subunits that are essential for the catalytic functions of the enzyme. Most of the active site labeling of the enzyme has been carried out at the substrate level. The only subunit that has been studied, with the help of fluorescence spectroscopy in some detail, is the regulatory subunit of E. coli RNA polymerase or the subunit.