title: Six missense mutations associated with type I and type II protein C deficiency and implications obtained from molecular modelling.
abstract: The molecular basis of protein C deficiency was studied in three type I and three type II heterozygotes. Three probands showed thrombotic complications. All the exons and intron/exon junctions of the protein C gene were studied using a strategy combining by the polymerase chain reaction (PCR) amplification, single-strand conformational polymorphism (SSCP) analysis, and DNA sequencing of the PCR-amplified fragments. Six missense mutations were identified, including three novel ones. One was located in exon II, in which the initiating translation codon (ATG) encoding for Met at position -42 was replaced by ACG encoding for Thr. The other five were located in exon IX, and included TAC(Tyr399)-->CAC(His), CCG(Pro327)-->CTG(Leu), GAC(Asp359)-->AAC(Asn) in two cases, and GGG(Gly350)-->AGG(Arg). Four of the six missense mutations occurred in CG dinucleotide. Sequence analysis of the other exons excluded additional mutations. By restriction enzyme analysis, co-segregation of the mutation with protein C deficiency was observed in four families. The other two mutations at amino acid positions -42 and 350 were also considered to be associated with protein C deficiency due to the absence of these mutations in 50 normal individuals. A structural model of the protease domain of mutant activated protein C was constructed by the chimeric modelling method, and the resultant model suggested conformational changes due to each missense mutation identified in protein C deficiency. The present data also provide some evidence regarding the genetic heterogeneity of protein C deficiency.
authors.affiliation: Laboratory of Thrombosis Research, National Cardiovascular Center Research Institute, Suita, Japan.
authors.forename: Y Z
authors.initials: YZ
authors.lastname: Zheng
authors.forename: T
authors.initials: T
authors.lastname: Sakata
authors.forename: T
authors.initials: T
authors.lastname: Matsusue
authors.forename: H
authors.initials: H
authors.lastname: Umeyama
authors.forename: H
authors.initials: H
authors.lastname: Kato
authors.forename: T
authors.initials: T
authors.lastname: Miyata
mesh_terms: D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D001483:Base Sequence; D003062:Codon; D004262:DNA Restriction Enzymes; D005091:Exons; D005260:Female; D006801:Humans; D007438:Introns; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010375:Pedigree; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D011486:Protein C / Q000737:chemistry / Q000235:genetics*; D020151:Protein C Deficiency; D017422:Sequence Analysis, DNA; D013927:Thrombosis / Q000201:enzymology / Q000235:genetics
chemical_list: D003062:Codon; D011486:Protein C; D004262:DNA Restriction Enzymes