Physicochemical studies of pregnancy-specific .beta.1-glycoprotein; unusual ultracentrifugal and circular dichroic properties

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Abstract
The molecular properties of native and modified pregnancy-specific .beta.1-glycoprotein (SP1) from human placenta were evaluated by sedimentation equilibrium, gel electrophoresis and circular dichroic measurements. Native SP1 contained 6.2% N-acetylneuraminic acid (NANA), 5.8% galactose (Gal), 13% N-acetylglucosamine, 6.5% mannose and 1.1% fucose but no detectable N-acetylgalactosamine. Treatment with mixed exoglycosidases and .alpha.-mannosidase removed 79% of the carbohydrate including all of the NANA and Gal. The intensity of the circular dichroic spectrum of SP1 in the far UV was quite low with a positive maximum at 235 nm and a negative maximum at 215 nm. The 235-nm band was lost upon treatment with reducing agents or with guanidinium chloride (GdmCl), but not by treatment with neuraminidase. Treatment of SP1 with neuraminidase, or with mixed exoglycosidases and .alpha.-mannosidase, resulted in decreases of the apparent MW obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Neither exposure of SP1 to GdmCl nor its reduction and alkylation resulted in the appearance of subunits on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The partial specific volume of SP1 determined experimentally by comparing sedimentation equilibrium profiles in H2O and D2O was 0.695 .+-. 0.007 ml/g. The MW of SP1 in 6 M GdmCl (in the presence or absence of reducing agents) by equilibrium sedimentation was 42,300 .+-. 400. In the absence of denaturing agents, SP1 existed in the form of aggregates (at least as high as trimeric SP1) that dissociated only slowly upon dilution. The presence of these aggregates may contribute to the reported molecular heterogeneity of SP1.