Isolation of Fibrinogen Protomer by Partial Reduction of Disulfide Bonds in Concentrated Urea

A Albert, F Cacho, J Saiz
1979 VIIth International Congress on Thrombosis and Haemostasis   unpublished
Partial reduction of human and bovine fibrinogen (F). recently dissolved in 6 and 8M deionized urea, with different amounts of dithioeritritol (DTE) (10 to 60:100 DTE:F, w/w) has been carried out in nitrogen atmosphere. Disc gel electrophoresis in 0.1% SDS shows changing amounts of six molecular species depending on the reaction time and DTE:F ratio. Three bands are identical with those obtained by total reduction of fibrinogen, one with unreduced material, the remaining two show mobilities
more » ... esponding to molecular weights of 170.000-190.000 (band H, with yields up to 40-50%) and 130.000-140.000 (band N).Total reduction of the isolated species H shows identical composition in fibrinogen chains as the original material. Stabilization by alkylation of this soecies is only successful in 0.1% SDS. Comparative ultracentrifugation studies show sedimentation coefficients of 6.8 S for fibrinogen incubated with SDS and 4.1 S for the H Species, which agree with a ratio 2:1 for their molecular weights.These results confirm the existence of two protomer entities easily cleaved by partial reduction but the need of high concentration of urea to separate the protomer, suggests the existence of additional strong non-covalent interactions between the two hemimolecules.
doi:10.1055/s-0038-1665773 fatcat:yof2w5k5wbcgbh5c2ocwkcboza