Diagnosis of Inherited Disorders of Galactose Metabolism

Abstract

Galactose metabolism occurs through an evolutionarily conserved pathway in which galactose and uridine diphosphoglucose are converted to glucose-1-phosphate and uridine diphosphogalactose through the action of three sequential enzymes: galactokinase (GALK, EC 2.7.1.6), galactose-1-phosphate uridyltransferase (GALT, EC 2.7.7.12), and uridine phosphogalactose 4'-epimerase (GALE, EC 5.1.3.2). Inborn errors of galactose metabolism occur with impaired activity for each of the enzymes. Classical galactosemia is the most common and the most severe of these diseases and is caused by deficiency of the GALT enzyme, affecting from approximately 1 in 10,000 to 1 in 30,000 live births. Deficiency of GALE is the rarest of the three diseases. Assays for galactitol and galactose-1-phosphate and methods for assaying enzyme activities of GALT, GALK, and GALE are provided here. Interpretation of diagnostic results for screen-positive newborns or symptomatic patients, as well as therapeutic interventions based on biochemical phenotype and molecular genotype, are also included as decision trees.

Metabolic and Phenotypic Characterization of Human Skin Fibroblasts After Forcing Oxidative Capacity

“…Galactose in OXPHOSm must be phosphorylated by galactokinase 1, resulting in galactose-1-phosphate. Galactose-1- phosphate is then converted into glucose-1-phosphate by uridyl transferase, which uses UDP-glucose (Cuthbert et al, 2008). UDP-galactose-4-epimerase catalyzes the reversible conversion of UDP-galactose to UDP-glucose in galactose utilization metabolism (Cuthbert et al, 2008).…”

“…Galactose-1- phosphate is then converted into glucose-1-phosphate by uridyl transferase, which uses UDP-glucose (Cuthbert et al, 2008). UDP-galactose-4-epimerase catalyzes the reversible conversion of UDP-galactose to UDP-glucose in galactose utilization metabolism (Cuthbert et al, 2008). With this work, we demonstrated that the mitochondrial remodeling promoted by OXPHOSm in human skin fibroblasts increased UDP-galactose-4-epimerase (GALE) gene expression ( Figure 7G), possibly as an adaptive response due to the modified cell culture medium used.…”

Quantification of Galactose-1-Phosphate Uridyltransferase Enzyme Activity by Liquid Chromatography–Tandem Mass Spectrometry

“…The diagnosis of classical galactosemia usually involves the measurement of GALT enzyme activity in erythrocytes, and the enzyme activity is usually absent or barely detectable. Several fluorescent and radioactive enzyme assays for GALT have been described (4–11) since the causative enzyme defect was identified in 1956 in the Kalckar laboratory (12). Unfortunately, these assays are usually nonspecific and laborious, and do not demonstrate sufficient performance in measuring low enzyme activity (<5% of control values).…”

Multiplex Enzyme Assay for Galactosemia Using Ultraperformance Liquid Chromatography–Tandem Mass Spectrometry

“…For babies with positive screening results, i.e., increased galactose and/or galactose-1-phosphate, confirmatory analyses should follow. These should include quantitative enzyme assays from fresh whole blood, galactose-1-phosphate analysis in erythrocytes, and mutational analysis of galactokinase 1 (GALK1) 6 , UDP-galactose-4-epimerase (GALE), and galactose-1-phosphate uridylyltransferase (GALT) genes (7 ). Numerous disease-causing mutations have been identified, more than 200 mutations in GALT, 31 mutations in GALK1, and 20 mutations in GALE (8 ).…”