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Fibrinogen Kaiserslautern (γ 380 Lys to Asn): a new glycosylated fibrinogen variant with delayed polymerization
Author(s) -
Ridgway Hayley J.,
Brennan Stephen O.,
Loreth Ralph M.,
George Peter M.
Publication year - 1997
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1046/j.1365-2141.1997.4363246.x
Subject(s) - sialic acid , glycosylation , biochemistry , chemistry , oligosaccharide , fibrinogen , thrombin , asparagine , microbiology and biotechnology , biology , amino acid , platelet , immunology
An adult woman diagnosed with cerebral thrombosis following a caesarean section was found to have severely prolonged thrombin and reptilase times. Five other family members also had prolonged, but variable, thrombin and reptilase times. Analysis of purified fibrinogen on reducing SDS‐PAGE revealed an additional band, in all family members, which migrated immediately below the normal Bβ band. Western blotting indicated that this band was a gamma chain and endoglycosidase‐F digestion established that it contained an additional oligosaccharide side chain. Partial acid hydrolysis localized the new oligosaccharide to the C‐terminus of the gamma chain. Amplification of this region by PCR and subsequent DNA sequencing demonstrated a single base substitution altering the normal 380 Lys (AAG) codon to Asn (AAT), producing a new Asn‐Lys‐Thr glycosylation site. The propositus and one other family member were homozygous for this mutation but the remaining four family members were heterozygous. The polymerization of purified fibrin monomers from the propositus was grossly abnormal; however, the polymerization curve was almost normalized by the removal of terminal sialic acid residues. This suggests that the polymerization defect was primarily caused by additional negatively charged sialic acid residues present on the new oligosaccharide. Further analysis of the D domain of purified fibrinogen established that calcium binding to the high affinity site remained unaffected by the bulky carbohydrate side chain or negatively charged sialic acid residues.