Abstract
Glutaric acidemia type 1 is an inherited metabolic disorder which can cause macrocephaly, muscular rigidity, spastic paralysis and other progressive movement disorders in humans. The defects in glutaryl-CoA dehydrogenase (GCDH) associated with this disease are thought to increase holoenzyme instability and reduce cofactor binding. Here, the first structural analysis of a GCDH enzyme inthe absence of the cofactor flavin adenine dinucleotide (FAD) is reported. Theapo structure of GCDH from Burkholderia pseudomallei reveals a loss of secondary structure and increased disorder in the FAD-binding pocket relative to the ternary complex of the highly homologous human GCDH. After conducting a fragment-based screen, four small molecules were identified which bind to GCDH from B. pseudomallei. Complex structures were determined for these fragments, which cause backbone and side-chain perturbations to key active-site residues. Structural insights from this investigation highlight differences from apo GCDH and the utility of small-molecular fragments as chemical probes for capturing alternative conformational states of preformed protein crystals.
Highlights
Glutaryl-CoA dehydrogenase (GCDH) is an acyl-CoA dehydrogenase (ACDH) which catalyzes an intermediate step in the metabolic breakdown of lysine and tryptophan (Gomes et al, 1981; Kim & Miura, 2004; Lenich & Goodman, 1986; Goodman et al, 1995)
Defects in glutaryl-CoA dehydrogenase (GCDH) are reponsible for glutaric acidemia type 1 (GA1), an inherited metabolic disorder which prevents the complete breakdown of lysine and tryptophan (Strauss et al, 2003; Strauss & Morton, 2003; Goodman et al, 1998)
We have determined the first structure of an intact GCDH holoenzyme without flavin adenine dinucleotide (FAD) or any small molecules present in either the substrate- or cofactor-binding pockets
Summary
Glutaryl-CoA dehydrogenase (GCDH) is an acyl-CoA dehydrogenase (ACDH) which catalyzes an intermediate step in the metabolic breakdown of lysine and tryptophan (Gomes et al, 1981; Kim & Miura, 2004; Lenich & Goodman, 1986; Goodman et al, 1995). Defects in GCDH are reponsible for glutaric acidemia type 1 (GA1), an inherited metabolic disorder which prevents the complete breakdown of lysine and tryptophan (Strauss et al, 2003; Strauss & Morton, 2003; Goodman et al, 1998). This disease manifests as macrocephaly and bleeding abnormalities in newborns, as well as muscular rigidity, spastic paralysis and other progressive movement disorders in older individuals (Strauss et al, 2003; Hedlund et al, 2006). Controversy persists concerning attempts to connect phenotype to specific genotypic markers of the disease, as patients symptomatic for GA1 sometimes do not present with GCDH deficiency (Westover et al, 2003; Christensen et al, 2004; Garcia et al, 2008)
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