[Molecular analysis of the structure of the mutant NADH-cytochrome b5 reductase gene causing methemoglobinemia].

Hereditary methemoglobinemia is a genetic disorder, transmitted as an autosomal recessive trait, characterized by NADH-cytochrome b5 reductase (b5R) deficiency. It is classified three types, erythrocyte, generalized, and blood cell types, in terms of tissues in which the enzyme is deficient. In attempt to analyse molecular mechanisms involved in the enzyme deficiency, we isolated b5R genes from the patient of hereditary methemoglobinemia, generalized type. 1. Sequence comparison of both b5R genes indicated that a thymidine at first position of codon 127 was altered to a cytidine, resulting in replacement of serine with proline. 2. Dot blot hybridization of the PCR-amplified DNA with allele-specific oligonucleotide probes showed that the patient was homozygous for this mutation. 3. Secondary structure prediction by the Chou-Fasman method showed that Ser-127 is presumed to be in an alpha-helix structure of a nucleotide-binding domain. 4. These observations showed that extension of the alpha-helix structure caused by replacement of Pro-127 affects the electron transport and the result is a generalized type of hereditary methemoglobinemia.