Dynamic Fluctuations of Protein-Carbohydrate Interactions Promote Protein Aggregation
Figure 2
Peptide-carbohydrate interactions are disrupted upon heat stress.
(A) a full-antibody cartoon illustrates the positions of the variants with engineered surface cysteines, and labeling approach in this set of experiments. Carbohydrates were labeled with Alexa350 hydrazide dye. Surface cysteines for each variant were labeled with Alexa488 maleimide dye. Variants 2 and 1 have surface cysteines engineered in the CH2 domain, and Variants 4 and 3 in the Fab region. (B) Ratiometric analysis of fluorescence data for double-labeled samples after heat stress at 60°C for 2 hours. Acceptor/donor ratios of stressed (2 hrs) and unstressed (0 hrs) samples are shown. The number above each pair of columns indicates the change in distance between the donor and acceptor fluorophores in the initial and final state. (C) Steady-state fluorescence data for individual double-labeled variants.