UDP-glucose Dehydrogenase Polymorphisms from Patients with Congenital Heart Valve Defects Disrupt Enzyme Stability and Quaternary Assembly

Abstract

Background: UDP-glucose dehydrogenase (UGDH) polymorphisms were identified in a screen of candidate genes for heart valve defects. Results: Two individual mutants fail to rescue cardiac valve defects in UGDH-deleted zebrafish and have reduced stability in vitro. Conclusion: UGDH loss of function mutations result in a subset of human congenital cardiac valve defects caused by reduced enzyme activity during morphogenesis. Significance: Screening these alleles could predict valve defects.

UDP-glucose Dehydrogenase Activity and Optimal Downstream Cellular Function Require Dynamic Reorganization at the Dimer-Dimer Subunit Interfaces

“…A thermal denaturation assay was previously used to show that the melting temperature (T m ) of the UGDH apoenzyme was increased by 20°C in the presence of saturating substrate and cofactor to form the holoenzyme (4,29). We extended our prior characterization of the wild-type enzyme using this assay in the presence of either reduced or oxidized cofactor alone, substrate or product alone, and each permutation of the ternary complex (e.g.…”

“…UGDH targeted disruption and loss of function polymorphisms that interfere with the enzyme quaternary structure and cellular stability have been implicated in cardiac malformation in multiple organisms (4,5,33,34). Moreover, the androgenstimulated expression of UGDH is important for prostate epithelial androgen solubilization and elimination.…”

“…In prostate tumor cell lines, UGDH is a constitutively expressed but androgen-stimulated gene product, up-regulation of which can specifically drive excess steroid elimination through glucuronidation in hormone-dependent cells (12). In the developing heart, UGDH point mutations that alter the integrity of quaternary structure, normally hexameric, fail to support maximal HA production that is needed for cardiac valve formation (4). Clinical and cellular consequences associated with UGDH defective function implicate stable but dynamic association of subunits and suggest useful therapeutic applications would focus on sustaining robust UGDH activity.…”

“…In all cases the enzyme oxidizes UDP-glucose through two NAD ϩ -dependent electron transfers. Of particular interest to studies relating quaternary structure to cellular function is that bacterial UGDH adopts a strictly dimeric conformation in the presence or absence of its substrate and cofactor (27), whereas higher eukaryotic UGDH orthologues purify in a hexamer-dimer equilibrium that is stabilized to hexameric association under conditions representative of the cellular environment (4,11,16,28). Kinetic data comparing wild-type UGDH to an obligate (inactive) hexamer mutant in which residue 132 was deleted (25) together with the interesting observation of an "open" conformation of the wild-type enzyme observed in cocrystals with the potent UGDH inhibitor UDP-xylose suggest that the transient capacity to dissociate and reorganize the hydrogen bond network at the interface between dimeric units is an important element of the normal catalytic cycle.…”

“…In adult tissues, in wound healing, and during development, high levels of UDP-glucuronate are used to synthesize extracellular glycosaminoglycans such as heparan sulfate and hyaluronan (HA) 4 (2), and the decarboxylation of UDP-glucuronate to UDP-xylose initiates proteoglycan production in the Golgi (3). In embryonic development, temporal and spatial elevation in UDP-glucuronate synthesis are indispensible for rapid extrusion of HA to the extracellular space to effect the dramatic morphological changes that generate anatomical structures such as cardiac valves (4,5).…”