Defective fibrinogen polymerization associated with a novel gamma279Ala-->Asp mutation

A woman with menorrhagia was investigated for a suspected fibrinogen mutation when coagulation tests revealed prolonged thrombin (55 s) and reptilase (43 s) times together with a functional and an antigenic fibrinogen concentration of 0.7 and 2.8 mg/ml respectively. Heterozygosity for a gamma-chain mutation was suggested by a doublet gamma band on SDS-PAGE and an increased negative charge was observed on isoelectric focusing of HPLC-isolated gamma-chains. Electrospray ionization mass spectrometry revealed a gamma-chain mass of 48 411 Da, which was 20 Da more than the control value of 48 391 Da. Because the normal and variant gamma-chains were not resolved, this implied a 40-Da increase in 50% of the gamma-molecules. An increased negative charge and a 44-Da increase in mass was verified when DNA sequencing showed heterozygosity for an Ala (GCC)-->Asp (GAC) substitution at codon 279 of the gamma-gene. Fibrin polymerization curves indicated a delay in the onset, and a decrease in the rate, of polymerization. Examination of crystal structures showed that the adjacent Tyr-gamma280 side chain is involved in bonding across the D-D interface, and from the proximity of the gamma279Ala-->Asp mutation it would appear that this perturbs the end-to-end DD interactions between fibrin units of the growing polymer.