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Title: An active site–tail interaction in the structure of hexahistidine-tagged Thermoplasma acidophilum citrate synthase

Citrate synthase from the thermophilic euryarchaeon T. acidophilum fused to a hexahistidine tag was purified and biochemically characterized. The structure of the unliganded enzyme at 2.2 Å resolution contains tail–active site contacts in half of the active sites. Citrate synthase (CS) plays a central metabolic role in aerobes and many other organisms. The CS reaction comprises two half-reactions: a Claisen aldol condensation of acetyl-CoA (AcCoA) and oxaloacetate (OAA) that forms citryl-CoA (CitCoA), and CitCoA hydrolysis. Protein conformational changes that ‘close’ the active site play an important role in the assembly of a catalytically competent condensation active site. CS from the thermoacidophile Thermoplasma acidophilum (TpCS) possesses an endogenous Trp fluorophore that can be used to monitor the condensation reaction. The 2.2 Å resolution crystal structure of TpCS fused to a C-terminal hexahistidine tag (TpCSH6) reported here is an ‘open’ structure that, when compared with several liganded TpCS structures, helps to define a complete path for active-site closure. One active site in each dimer binds a neighboring His tag, the first nonsubstrate ligand known to occupy both the AcCoA and OAA binding sites. Solution data collectively suggest that this fortuitous interaction is stabilized by the crystalline lattice. As a polar but almostmore » neutral ligand, the active site–tail interaction provides a new starting point for the design of bisubstrate-analog inhibitors of CS.« less
Authors:
; ;  [1]
  1. Purdue University, 175 South University Street, West Lafayette, IN 47907-2063 (United States)
Publication Date:
OSTI Identifier:
22420144
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section F, Structural Biology Communications; Journal Volume: 71; Journal Issue: Pt 10; Other Information: PMCID: PMC4601594; PMID: 26457521; PUBLISHER-ID: ub5080; PUBLISHER-ID: S2053230X15015939; OAI: oai:pubmedcentral.nih.gov:4601594; Copyright (c) Murphy et al. 2015; This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CITRATES; CRYSTAL STRUCTURE; INTERACTIONS; LIGANDS; RESOLUTION