Sulfite oxidase (SO) deficiency, an inherited disease that causes severe neonatal neurological problems and early death, arises from defects in the biosynthesis of the molybdenum cofactor (Moco) (general sulfite oxidase deficiency) or from inborn errors in the SUOX gene for SO (isolated sulfite oxidase deficiency, ISOD). The X-ray structure of the highly homologous homonuclear dimeric chicken sulfite oxidase (cSO) provides a template for locating ISOD mutation sites in human sulfite oxidase (hSO). Catalysis occurs within an individual subunit of hSO, but mutations that disrupt the hSO dimer are pathological. The catalytic cycle of SO involves five metal oxidation states (MoVI, MoV, MoIV, FeIII, FeII), two intramolecular electron transfer (IET) steps, and couples a two-electron oxygen atom transfer reaction at the Mo center with two one-electron transfers from the integral b-type heme to exogenous cytochrome c, the physiological oxidant. Several ISOD examples are analyzed using steady-state, stopped-flow, and laser flash photolysis kinetics and physical measurements of recombinant variants of hSO and native cSO. In the structure of cSO, Mo…Fe = 32 Å, much too long for efficient IET through the protein. Interdomain motion that brings the Mo and heme centers closer together to facilitate IET is supported indirectly by decreasing the length of the interdomain tether, by changes in the charges of surface residues of the Mo and heme domains, as well as by preliminary molecular dynamics calculations. However, direct dynamic measurements of interdomain motion are in their infancy.

Molybdenum cofactor deficiency (MoCD) includes three ultrarare autosomal recessive inborn errors of metabolism (MoCD type A [MoCD-A], MoCD-B, and MoCD-C) that cause sulfite intoxication disorders. This natural history study analyzed retrospective data for 58 living or deceased patients (MoCD-A, n = 41; MoCD-B, n = 17). MoCD genotype, survival, neuroimaging, and medical history were assessed retrospectively. Prospective biomarker data were collected for 21 living MoCD patients. The primary endpoint was survival to 1 year of age in MoCD-A patients. Of the 58 MoCD patients, 49 (MoCD-A, n = 36; MoCD-B, n = 13) had first presenting symptoms by Day 28 (neonatal onset; median: 2 and 4 days, respectively). One-year survival rates were 77.4% (overall), 71.8% (neonatal onset MoCD-A), and 76.9% (neonatal onset MoCD-B); median ages at death were 2.4, 2.4, and 2.2 years, respectively. The most common presenting symptoms in the overall population were seizures (60.3%) and feeding difficulties (53.4%). Sequelae included profound developmental delay, truncal hypotonia, limb hypertonia that evolved to spastic quadriplegia or diplegia, dysmorphic features, and acquired microcephaly. In MoCD-A and MoCD-B, plasma and urinary xanthine and S-sulfocysteine concentrations were high; urate remained below the normal reference range. MOCS1 mutation homozygosity was common. Six novel mutations were identified. MoCD is a severe neurodegenerative disorder that often manifests during the neonatal period with intractable seizures and feeding difficulties, with rapidly progressive significant neurologic disabilities and high 1-year mortality rates. Delineation of MoCD natural history supports evaluations of emerging replacement therapy with cPMP for MoCD-A, which may modify disease course for affected individuals.

Mammalian sulfite oxidase (SO) is a dimeric enzyme consisting of a molybdenum cofactor- (Moco) and haem-containing domain and catalyses the oxidation of toxic sulfite to sulfate. Following sulfite oxidation, electrons are passed from Moco via the haem cofactor to cytochrome c, the terminal electron acceptor. In contrast, plant SO (PSO) lacks the haem domain and electrons shuttle from Moco to molecular oxygen. Given the high similarity between plant and mammalian SO Moco domains, factors that determine the reactivity of PSO towards oxygen, remained unknown. In the present study, we generated mammalian haem-deficient and truncated SO variants and demonstrated their oxygen reactivity by hydrogen peroxide formation and oxygen-consumption studies. We found that intramolecular electron transfer between Moco and haem showed an inverse correlation to SO oxygen reactivity. Haem-deficient SO variants exhibited oxygen-dependent sulfite oxidation similar to PSO, which was confirmed further using haem-deficient human SO in a cell-based assay. This finding suggests the possibility to use oxygen-reactive SO variants in sulfite detoxification, as the loss of SO activity is causing severe neurodegeneration. Therefore we evaluated the potential use of PEG attachment (PEGylation) as a modification method for future enzyme substitution therapies using oxygen-reactive SO variants, which might use blood-dissolved oxygen as the electron acceptor. PEGylation has been shown to increase the half-life of other therapeutic proteins. PEGylation resulted in the modification of up to eight surface-exposed lysine residues of SO, an increased conformational stability and similar kinetic properties compared with wild-type SO.