Chromatographic, ultracentrifugal, and related studies of fibrinogen “Baltimore”

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Abstract
Chromatographic, ultracentrifugal, and related studies of the fibrinogen of a patient with a congenital disorder of fibrinogen (fibrinogen “Baltimore” have provided evidence of structural differences from normal. Diethylaminoethyl-cellulose (DEAE-cellulose) gradient elution chromatography demonstrated two major peaks in the elution pattern of fibrinogen Baltimore as was the case for normal fibrinogen. However, the first peak of fibrinogen Baltimore was somewhat broader and more symmetrical and was eluted significantly later in the chromatogram than the corresponding peak of normal fibrinogen. Additionally, in some elution patterns, a shoulder on the ascending limb of peak 1 was present, suggesting the presence of chromatographically “normal” fibrinogen. Thrombin time determinations of eluted column fractions from a chromatogram of propositus fibrinogen supported this conclusion by demonstrating that fibrinogen from the ascending portion of peak 1 behaved functionally more like normal than that later in the chromatogram. Chromatograms of mixtures of propositus and normal fibrinogen confirmed the ability of this technique to distinguish normal from Baltimore fibrinogen. Chromatograms of fibrinogen isolated from two affected daughters displayed the characteristic increased anionic binding of peak 1 fibrinogen. Sedimentation velocity experiments indicated that the So20, [unk] of fibrinogen Baltimore was slightly greater (8.13S vs. 7.85S) than that of normal fraction I-4. Differences in concentration dependence (- 0.65 c vs. - 1.30 c for normal) of the sedimentation coefficient could be attributable in part to spatial conformational differences. Molecular sieving experiments in acrylamide gels indicated that the molecular weight of propositus fraction I-2 was about the same as that of normal fibrinogen of comparable solubility (i.e. I-4, mol wt 325,000). Studies of the UV spectra, tyrosine/tryptophan ratios, sialic acid and hexose content, and N-terminal amino acids demonstrated no consistent significant differences from normal fraction I-4.