Analysis of the Glycosylation Profile of Disease-Associated Water-Soluble Prion Protein Using Lectins

Figure 3

Glycosylation profile of hamster plasma total proteins is essential for their stability in blood. ((a)-(c)) Binding profile of ConA, Concanavalin A; WGA, wheatgerm agglutinin; RCA, Ricinus communis agglutinin; DBA, Dolichos biflorus agglutinin; PNA, peanut agglutinin; SBA, soybean agglutinin; and UEA-1, Ulex europaeus agglutinin I to equal amounts of total proteins from noninfected and terminally scrapie-infected hamster plasma ((a) and (b)) and from scrapie-infected hamster plasma collected at different time points after infection (c). ((a) and (c)) Lectin-ELISA analysis, (b) lectin-blot analysis (10% SDS-PAGE gel). The figures show that lectins binding profile to plasma total proteins is not due to the infection or disease duration but rather to potential relevance of the glycans recognized by ConA and RCA for maintaining the stability of plasma total proteins including PrP against certain insults. (d) Comparison between the regression lines of the binding profile of ConA, WGA, RCA, DBA, PNA, SBA, and UEA-1 toward plasma total proteins throughout disease period. The basal immunoreactivity of ConA and RCA is the highest and this trend is maintained also throughout disease period; only immunoreactivity of ConA (increase) and DBA (decrease) correlates with disease progression (r = 0.90); lectins basal immunoreactivity values are potential predictors of the expected effect by the relative sugars on proteins stability state. (e) The increase in the immunoreactivity of ConA is proportional to the sum of reduction in the immunoreactivity of DBA, SBA, and UEA-I. The left side of the figure shows that the two effects (decrease and increase of immunoreactivity) are inversely correlated between each other during disease progression, while the right side of the figure shows that the two effects (decrease and increase of immunoreactivity) are mathematically very close but opposite. The increase and decrease in the lectins reactivity were calculated by normalization of the absorbance values against the basal reactivity values for each lectin. Lectins basal reactivity is indicated by a circle shown on Figure 3(d). The sizes of molecular mass markers in kilodaltons are indicated on the left. Data are means ± SD and are representative of at least three independent assays and three different preparations, performed in duplicate. P<0.05, P<0.01.