UBC Theses and Dissertations
Comparison of the carbohydrate composition of α₂-macroglobulin from controls and patients with cystic fibrosis Park, Carol Maylene
The putative involvement of α₂-macroglobulin (α₂M) in the pathogenesis of Cystic Fibrosis (CF) has long been a subject of controversy. Previous reports have indicated that there are alterations in the carbohydrate moieties of α₂M. The objective of the present study was to compare the carbohydrate composition of α₂M isolated from the plasma of patients with CF and from the plasma of age- and sex-matched normal controls. The gas-liquid chromatographic procedure of Lehnhardt and Winzler was investigated using both α₂M and α₁-acid glycoprotein and modified to obtain the required sensitivity, precision and accuracy for these analyses. Investigation of the hydrolysis of both sugar mixtures and glycoproteins showed that significant losses of the released sugars occurred with extended hydrolysis and that use of the recommended hydrolysis times led to underestimation of the absolute carbohydrate composition. Analysis of the rates of degradation of mixtures of free sugars indicated that not all are degraded at the same rate. This implies that estimates of glycoprotein composition based on molar ratios are also in error. Therefore, hydrolysis times were individually determined for the two glycoproteins studied; those chosen were a compromise between maximal " release and minimal destruction of the various carbohydrate components. In contrast to the hydrolysis time recommended by Lehnhardt and Winzler (40 hr for α₁-acid glycoprotein), the hydrolysis times selected for α₁-acid glycoprotein and α₂M, were 30 hr and 35 hr, respectively. The analysis of α₂M from six CF patients and their age- and sex-matched normal controls using the modified gas-liquid chromatographic procedure indicated that there were no significant differences between the carbohydrate compositions of the glycoproteins. The carbohydrate composition of CF α₂M expressed as mean ± S.D. μmole carbohydrate per 100 mg protein was: fucose, 0.70 ± 0.12; mannose, 14.07 ± 1.31;. galactose, 6.72 ± 0.65; glucosamine, 15.38 + 1.59; sialic acid, 5.52 ± 0.33 and that of normal control α₂M was: fucose, 0.69 ± 0.11; mannose, 14.42 ± 1.21; galactose, 6.91 ± 0.52; glucosamine, 16.13 ± 1.77; sialic acid, 5.58 ± 0.31. Therefore, contrary to previous reports, this thesis demonstrates that there is no difference in the carbohydrate composition of α₂M in CF.