CHARACTERIZATION OF DIGOXIGENIN DERIVATIVES OF HUMAN COAGULATION-FACTOR-VIIA AND COAGULATION-FACTOR-X BY ELECTROSPRAY MASS-SPECTROMETRY ANDENZYME-KINETICS
Sv. Thiruvikraman et al., CHARACTERIZATION OF DIGOXIGENIN DERIVATIVES OF HUMAN COAGULATION-FACTOR-VIIA AND COAGULATION-FACTOR-X BY ELECTROSPRAY MASS-SPECTROMETRY ANDENZYME-KINETICS, Rapid communications in mass spectrometry, 10(11), 1996, pp. 1367-1370
Digoxigenin ester (Dig) derivatives of coagulation factors VIIa and X
facilitate staining studies to localize tissue factor activity in huma
n atherosclerotic plaques. The larger the number of attached Dig units
the easier it is to form highly visual stains. Electrospray ionizatio
n was used to characterize the Dig derivatives, as a function of exces
s derivatization reagent, to determine the optimal derivatives, i.e. t
he highest number of Dig units attached with the product still retaini
ng enzymatic activity. The enzymatic activity of factor VIIa derivatiz
ed at 50-fold Dig excess (with 28, 34, 48 or 52 Dig units attached, ma
sses to 79 kDa) remained the same as that of native factor VIIa. In co
ntrast, the enzymatic activity of factor X derivatives diminished abov
e 15-fold Dig excess (15 Dig units attached, mass 65.6 kDa). The distr
ibution of derivatized lysine, histidine, arginine, tryptophan and tyr
osine residues was estimated for several possible configurations.