Relationship between kinetic properties of mutant enzyme and biochemical and clinical responsiveness to biotin in holocarboxylase synthetase deficiency

Holocarboxylase synthetase (HCS) deficiency is a metabolic disorder that ca uses a biotin-responsive multiple carboxylase deficiency. We analyzed the k inetic properties of seven mutant HCS proteins. Two of these enzymes harbor ed mutations within the putative biotin-binding region of HCS and showed el evated K-m values for biotin compared with that of the wild-type form (K-m mutant; Gly581Ser: 45 times, delThr610: 3 times). The remaining five mutati ons (Arg183Pro, Leu216Arg, Leu237Pro, Val333GIu, and Val363Asp) were locate d outside the biotin-binding region. The enzymes containing these mutations showed normal or low K-m values for biotin (non-K-m mutant). Symptoms of p atients who have the non-K-m mutants, as well as those of patients who have the K-m mutants, responded to biotin therapy. This is probably because the K-m value for biotin of normal HCS is higher than the physiologic concentr ation of biotin in human cells. The V-max values of all mutant HCS proteins were considerably decreased, but to a variable degree. The responsiveness to biotin supplementation of propionyl-CoA carboxylase activity in cultured cells bearing the mutations correlated well with the degree of reduction i n the V-max of HCS. Patients who have mutant HCS proteins with lower V-max showed poorer clinical and biochemical responses to biotin therapy. These o bservations suggest that the reduction of V-max is an essential factor for pathophysiology and prognosis of HCS deficiency under treatment with large amounts of biotin. The determination of HCS genotype can be valuable for ch aracterizing the clinical phenotype in HCS deficient patients.