Effect of the carbohydrate moiety on the secondary structure of beta 2-glycoprotein. I. Implications for the biosynthesis and folding of glycoproteins.
By use of six highly purified exoglycosidases with well-defined specificity, the oligosaccharide units of human plasma beta 2-glycoprotein I (beta 2I) were modified by sequential enzymatic degradation. The released monosaccharides (NeuAc, Gal, GlcNAc, and Man) were quantified, and the carbohydrate compositions of the resulting glycoprotein (gp) derivatives were determined. The gp was found to be both partially sialylated and galactosylated. These findings which are in agreement with earlier reports suggest that the carbohydrate moiety of beta 2I possesses more bi- than tri-antennas, probably three of the former and two of the latter carbohydrate units. Circular dichroic (CD) spectra of native beta 2I and its derivatives were measured in aqueous buffer and 2-chloroethanol (2-CE). Analysis of these spectra for elements of secondary structure showed beta 2I and most of the derivatives to contain predominantly beta-sheet and beta-turn structures. The lack of alpha-helical structures in aqueous buffer was noted. Removal of a large portion of the carbohydrate moiety did not alter the CD spectra or secondary structure of beta 2I in either aqueous buffer or in 2-CE. However, after enzymatic removal of approximately 96% of the carbohydrate moiety, large significant changes in the spectra and secondary structures were observed. In aqueous buffer a shift in the wavelength minimum occurred, accompanied by an increase in the magnitude of the molar ellipticity and the amount of beta-turn, with a reduction in random coil. One-third of the amino acids which were originally in random coil conformation assumed beta-turns after removal of 96% of the carbohydrate moiety.(ABSTRACT TRUNCATED AT 250 WORDS)
Walsh
M T
MT
Department of Biophysics, Boston University School of Medicine, Massachusetts 02118.
Watzlawick
H
H
Putnam
F W
FW
Schmid
K
K
Brossmer
R
R
D002240:Carbohydrate Sequence; D002942:Circular Dichroism; D006023:Glycoproteins / Q000096:biosynthesis / Q000648:ultrastructure*; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D053482:beta 2-Glycoprotein I
D006023:Glycoproteins; D053482:beta 2-Glycoprotein I; D006026:Glycoside Hydrolases