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. 2021 Jul 26;9:707797. doi: 10.3389/fchem.2021.707797

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Copyright © 2021 Cooper, Bird, Sheng, Choi, Silkstone, Simons, Syrett, Piano, Ronda, Bettati, Paredi, Mozzarelli and Reeder.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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SDS-PAGE comparing A12 conjugation with maleimide-PEG and mono-sulfone PEG. Novex MW markers (M); Hb-A12 (1); Hb-A12 + 10 mM DTT 40°C 1 h incubation (2); maleimide-PEG-A12 reaction mixture (3); maleimide-PEG-A12 product (4); maleimide-PEG-A12 product +10 mM DTT 40°C + 1 h (5); mono-sulfone PEG-A12 reaction mixture (6); mono-sulfone PEG-A12 product (7); mono-sulfone PEG-A12 product +10 mM DTT 40°C + 1 h incubation (8). Stoichiometry for incubation: maleimide-PEG: Hb = 12:1; mono-sulfone-PEG: Hb = 3:1. Labels equate to % of total protein in lane in each band for: Di-PEGylated Hb monomer; Mono-PEGylated Hb Monomer; Hb dimer; Hb monomer.