Structural and energetic basis of isopropylmalate dehydrogenase enzyme catalysis. (4th October 2014)

• Record Type: Journal Article
• Authors: Palló, Anna; Oláh, Julianna; Gráczer, Éva; Merli, Angelo; Závodszky, Péter; Weiss, Manfred S.; Vas, Mária
• Is Part Of: FEBS journal. Volume 281:Number 22(2014)
• Page Start: 5063
• Page End: 5076
• Publication Date: 2014-10-04
• Subjects: Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pathology, Molecular -- Periodicals; 572
• DOI: 10.1111/febs.13044
• Languages: English
• ISSNs: 1742-464X

Abstract

<abstract abstract-type="main" id="febs13044-abs-0001"> <title> <x xml:space="preserve">Abstract</x> </title> <sec id="febs13044-sec-0001" sec-type="section"> <p>The three‐dimensional structure of the enzyme 3‐isopropylmalate dehydrogenase from the bacterium <italic>Thermus thermophilus</italic> in complex with Mn<sup>2+</sup>, its substrate isopropylmalate and its co‐factor product NADH at 2.0 Å resolution features a fully closed conformation of the enzyme. Upon closure of the two domains, the substrate and the co‐factor are brought into precise relative orientation and close proximity, with a distance between the C2 atom of the substrate and the C4N atom of the pyridine ring of the co‐factor of approximately 3.0 Å. The structure further shows binding of a K<sup>+</sup> ion close to the active site, and provides an explanation for its known activating effect. Hence, this structure is an excellent mimic for the enzymatically competent complex. Using high‐level QM/MM calculations, it may be demonstrated that, in the observed arrangement of the reactants, transfer of a hydride from the C2 atom of 3‐isopropylmalate to the C4N atom of the pyridine ring of NAD<sup>+</sup> is easily possible, with an activation energy of approximately 15 kcal·mol<sup>−1</sup>. The activation energy increases by approximately 4–6 kcal·mol<sup>−1</sup> when the K<sup>+</sup> ion is omitted from the calculations. In the most plausible scenario, prior to hydride transfer the ε‐amino group of Lys185 acts as a general base in the reaction, aiding the deprotonation reaction of 3‐isopropylmalate prior to hydride transfer by employing a low‐barrier proton shuttle mechanism involving a water molecule.</p> </sec> <sec id="febs13044-sec-0002" sec-type="section"> <title>Database</title> <p>Structural data have been submitted to the Protein Data Bank under accession number <ext-link ext-link-type="uri" xlink:href="http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4F7I" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">4F7I</ext-link>.</p> </sec> </abstract>