title: Hemophilia A due to mutations that create new N-glycosylation sites.
abstract: In studying the molecular defects responsible for cross-reacting material-positive hemophilia A, we have identified two patients in whom the nonfunctional factor VIII-like protein has abnormal, slower-moving heavy or light chains on SDS/PAGE. Both patients have severe hemophilia A (less than 1% of normal factor VIII activity) with a normal plasma level of factor VIII antigen. The molecular defects were identified by denaturing gradient gel electrophoresis screening of PCR-amplified products of the factor VIII-coding DNA sequence followed by nucleotide sequencing of the abnormal PCR products. In patient ARC-21, a methionine-to-threonine substitution at position 1772 in the factor VIII light chain creates a potential new N-glycosylation site at asparagine-1770. In patient ARC-22, an isoleucine-to-threonine substitution at position 566 creates a potential new N-glycosylation site at asparagine-564 in the A2 domain of the factor VIII heavy chain. The mobility of these chains on SDS/PAGE was normal after N-Glycanase digestion and procoagulant activity was generated--to a maximum of 23% and 45% of control normal plasma. Abnormal N-glycosylation, blocking factor VIII procoagulant activity, represents a newly recognized mechanism for the pathogenesis of severe hemophilia A.
authors.affiliation: Holland Laboratory, American Red Cross Blood Services, Rockville, MD 20855.
authors.forename: A M
authors.initials: AM
authors.lastname: Aly
authors.forename: M
authors.initials: M
authors.lastname: Higuchi
authors.forename: C K
authors.initials: CK
authors.lastname: Kasper
authors.forename: H H
authors.initials: HH
authors.lastname: Kazazian
authors.forename: S E
authors.initials: SE
authors.lastname: Antonarakis
authors.forename: L W
authors.initials: LW
authors.lastname: Hoyer
mesh_terms: D000581:Amidohydrolases / Q000494:pharmacology; D015153:Blotting, Western; D004252:DNA Mutational Analysis; D005169:Factor VIII / Q000235:genetics* / Q000378:metabolism; D006031:Glycosylation; D006467:Hemophilia A / Q000235:genetics*; D006801:Humans; D008970:Molecular Weight; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D013917:Thrombin / Q000378:metabolism
chemical_list: D005169:Factor VIII; D013917:Thrombin; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase