Visualizing an Allosteric Intermediate Using CuAAC Stabilization of an NMR Mixed Labeled Dimer.

ABSTRACT: Homodimers are the most abundant type of enzyme in cells, and as such, they represent the most elemental system for studying the phenomenon of allostery. In these systems, in which the allosteric features are manifest by the effect of the first binding event on a similar event at the second site, the most informative state is the asymmetric singly bound (lig₁) form, yet it tends to be thermodynamically elusive. Here we obtain milligram quantities of lig₁ of the allosteric homodimer, chorismate mutase, in the form of a mixed isotopically labeled dimer stabilized by Cu^(I)-catalyzed azide-alkyne cycloaddition (CuAAC) between the subunits. Below, we outline several critical steps required to generate high yields of both types of unnatural amino acid-containing

SUBMITTER: Sapienza PJ 

PROVIDER: S-EPMC9141113 | biostudies-literature | 2021 Dec

REPOSITORIES: biostudies-literature