A Plasma Fibrinogen Fraction With High Sialic Acid Content And Eecngated γ-Chains

Abstract

Having recently been able to separate Bβ- and γ-chains of human fibrinogen heterogeneous with respect to sialic acid, we were faced with the question whether a carbohydrate heterogeneity might be present in the original native fibrinoge.Using DEAE 52 cellulose and stepwise elution with phosphate buffer human fibrinogen from pooled or single donor plasma could indeed be separated into 3 populations with different sialic acid content. The most acidic fraction contained 30% more sialic acid (8 moles) than the original fibrinogen (6 moles). After reduction the 3 fibrinogen fractions were analyzed by two-dimensional electrophoresis. The first fraction (15%) contained partly degraded Aα-chains and a pattern of Bβ and γ-chains similar to the one described for the main fraction. The latter (70%) consisted of intact Act-chains, two main Bβ- and two main γ-chains and one additional minor band for each chain, all with the previously described carbohydrate heterogeneity.The third, most acidic fibrinogen fraction (15%) revealed an unusual γ-chain pattern. In addition to the two normal chains, two γ-chains with higher mol. wt. and more acidic IEP were found. The Aα- and Bβ-chains did not differ from those of the original fibrinogen. Again a heterogeneity with respect to sialic acid was visible both in the Bβ- and the normal as wsll as the elongated γ-chains. Desialylation of the elongated γ-chains resulted in only partial normalisation of the IEP.The binding of EDTA and citrate shows a strong ^-dependence suggesting a (partly) ionogenic binding between charged areas on the fibrinogen molecule and charged groups on EDTA or citrateIt has thus been possible to attribute the recently described elongated γ-chain variants to a particular sialic acid rich subfraction of normal human fibrinogen and therefore to conclude that the observed fibrinogen heterogeneity is at least to a considerable extent of inter- rather than intramolecular nature.