4HCX

Structure of ICDH-1 from M.tuberculosis complexed with NADPH & Mn2+

Summary for 4HCX

• Entry DOI: 10.2210/pdb4hcx/pdb
• Descriptor: Isocitrate dehydrogenase [NADP], MANGANESE (II) ION, NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE, ... (5 entities in total)
• Functional Keywords: icdh-1, rossmann fold, dehydrogenase, nadp binding, oxidoreductase
• Biological source: Mycobacterium tuberculosis
• Total number of polymer chains: 2
• Total formula weight: 92802.88

Authors

Hazra, S.,Blanchard, J. (deposition date: 2012-10-01, release date: 2013-02-27, Last modification date: 2024-02-28)

Primary citation

Quartararo, C.E.,Hazra, S.,Hadi, T.,Blanchard, J.S.
Structural, Kinetic and Chemical Mechanism of Isocitrate Dehydrogenase-1 from Mycobacterium tuberculosis.
Biochemistry, 52:1765-1775, 2013

PubMed Abstract: Mycobacterium tuberculosis (Mtb) is the leading cause of death due to a bacterial infection. The success of the Mtb pathogen has largely been attributed to the nonreplicating, persistence phase of the life cycle, for which the glyoxylate shunt is required. In Escherichia coli, flux through the shunt is controlled by regulation of isocitrate dehydrogenase (ICDH). In Mtb, the mechanism of regulation is unknown, and currently, there is no mechanistic or structural information about ICDH. We optimized expression and purification to a yield sufficiently high to perform the first detailed kinetic and structural studies of Mtb ICDH-1. A large solvent kinetic isotope effect [(D2O)V = 3.0 ± 0.2, and (D2O)(V/Kisocitrate) = 1.5 ± 0.3] and a smaller primary kinetic isotope effect [(D)V = 1.3 ± 0.1, and (D)(V/K[2R-(2)H]isocitrate) = 1.5 ± 0.2] allowed us to perform the first multiple kinetic isotope effect studies on any ICDH and suggest a chemical mechanism. In this mechanism, protonation of the enolate to form product α-ketoglutarate is the rate-limiting step. We report the first structure of Mtb ICDH-1 to 2.18 Å by X-ray crystallography with NADPH and Mn(2+) bound. It is a homodimer in which each subunit has a Rossmann fold, and a common top domain of interlocking β sheets. Mtb ICDH-1 is most structurally similar to the R132H mutant human ICDH found in glioblastomas. Similar to human R132H ICDH, Mtb ICDH-1 also catalyzes the formation of α-hydroxyglutarate. Our data suggest that regulation of Mtb ICDH-1 is novel.

PubMed: 23409873
DOI: 10.1021/bi400037w

Experimental method

X-RAY DIFFRACTION (2.18 Å)