Per-glycosylation of the Surface-Accessible Lysines: One-Pot Aqueous Route to Stabilized Proteins with Native Activity |
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Authors: | Raoul Walther Pere Monge Andreas Bøtker Pedersen Anja Benderoth Jannik Nedergaard Pedersen Azad Farzadfard Ole A. Mandrup Kenneth A. Howard Daniel E. Otzen Prof. Alexander N. Zelikin |
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Affiliation: | 1. Department of Chemistry, Aarhus University, Aarhus, 8000 Denmark;2. Interdisciplinary Nanoscience Centre (iNANO), Aarhus University, Aarhus, 8000 Denmark;3. Interdisciplinary Nanoscience Centre (iNANO), Aarhus University, Aarhus, 8000 Denmark Department of Molecular Biology and Genetics, Aarhus University, Aarhus, 8000 Denmark |
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Abstract: | Chemical glycosylation of proteins is a powerful tool applied widely in biomedicine and biotechnology. However, it is a challenging undertaking and typically relies on recombinant proteins and site-specific conjugations. The scope and utility of this nature-inspired methodology would be broadened tremendously by the advent of facile, scalable techniques in glycosylation, which are currently missing. In this work, we investigated a one-pot aqueous protocol to achieve indiscriminate, surface-wide glycosylation of the surface accessible amines (lysines and/or N-terminus). We reveal that this approach afforded minimal if any change in the protein activity and recognition events in biochemical and cell culture assays, but at the same time provided a significant benefit of stabilizing proteins against aggregation and fibrillation - as demonstrated on serum proteins (albumins and immunoglobulin G, IgG), an enzyme (uricase), and proteins involved in neurodegenerative disease (α-synuclein) and diabetes (insulin). Most importantly, this highly advantageous result was achieved via a one-pot aqueous protocol performed on native proteins, bypassing the use of complex chemical methodologies and recombinant proteins. |
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Keywords: | glucuronidation glycosylation insulin protein biochemistry synuclein |
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