Six members of a family with hypofibrinogenaemia had fibrinogen concentrations ranging from 0.5 to 1.1 mg/ml and, after sequencing the entire coding region and the intron exon boundaries of all three fibrinogen genes, a single heterozygous ACT-->ATT mutation was identified in the gamma gene. This novel mutation was not detected in normal family members or unrelated controls. The gamma371 Thr-->Ile substitution occurs at a conserved threonine in the gammaD domain, but molecules containing the new isoleucine were not present in circulating fibrinogen. The evidence for this was that purified gamma chains had a normal mass of 48375 Da compared to a control of 48374 Da, and tryptic peptide maps were entirely normal. The mutation predicts a mass increase of 12 Da in peptide T-36, but on mass mapping only the normal [M+2H] ion was detected, at 948 m/z. There was no new signal at 954 m/z that would indicate expression of variant chains. Also the normal 948 m/z signal was at the same intensity in digests from the proposita and controls. Crystal structures show a hydrogen bond from the threonine hydroxyl to the main chain and this case suggests this bond is critical in maintaining the structure of the gammaD domain.