Crystal Structure of Human Dihydrolipoamide Dehydrogenase: NAD[superscript +]/NADH Binding and the Structural Basis of Disease-causing Mutations

Journal Article · Tue Jul 13 04:00:00 EDT 2010 · J. Mol. Biol.

OSTI ID:1008644

Brautigam, Chad A; Chuang, Jacinta L; Tomchick, Diana R; Machius, Mischa; Chuang, David T ^[1]

---

1. U. of Texas-SMED

Human dihydrolipoamide dehydrogenase (hE3) is an enzymatic component common to the mitochondrial {alpha}-ketoacid dehydrogenase and glycine decarboxylase complexes. Mutations to this homodimeric flavoprotein cause the often-fatal human disease known as E3 deficiency. To catalyze the oxidation of dihydrolipoamide, hE3 uses two molecules: noncovalently bound FAD and a transiently bound substrate, NAD{sup +}. To address the catalytic mechanism of hE3 and the structural basis for E3 deficiency, the crystal structures of hE3 in the presence of NAD{sup +} or NADH have been determined at resolutions of 2.5 {angstrom} and 2.1 {angstrom}, respectively. Although the overall fold of the enzyme is similar to that of yeast E3, these two structures differ at two loops that protrude from the proteins and at their FAD-binding sites. The structure of oxidized hE3 with NAD{sup +} bound demonstrates that the nicotinamide moiety is not proximal to the FAD. When NADH is present, however, the nicotinamide base stacks directly on the isoalloxazine ring system of the FAD. This is the first time that this mechanistically requisite conformation of NAD{sup +} or NADH has been observed in E3 from any species. Because E3 structures were previously available only from unicellular organisms, speculations regarding the molecular mechanisms of E3 deficiency were based on homology models. The current hE3 structures show directly that the disease-causing mutations occur at three locations in the human enzyme: the dimer interface, the active site, and the FAD and NAD{sup +}-binding sites. The mechanisms by which these mutations impede the function of hE3 are discussed.

• Research Organization: Advanced Photon Source (APS), Argonne National Laboratory (ANL), Argonne, IL (US)
• Sponsoring Organization: USDOE
• OSTI ID: 1008644
• Journal Information: J. Mol. Biol., Journal Name: J. Mol. Biol. Journal Issue: (3) ; 2005 Vol. 350; ISSN JMOBAK; ISSN 0022-2836
• Country of Publication: United States
• Language: ENGLISH

Related Subjects

36 MATERIALS SCIENCE
CRYSTAL STRUCTURE
DECARBOXYLASES
DIMERS
DISEASES
ENZYMES
GLYCINE
ISOALLOXAZINES
MUTATIONS
NICOTINAMIDE
OXIDATION
OXIDOREDUCTASES
PROTEINS
YEASTS