Paper of the Month: January Edition
Thermal denaturation experiments for the characterization of protein stability are a well-established application of CD spectroscopy. In this study, these types of measurements drove a systematic approach to improve the thermal stability of VEGFR2-binding affibodies, that is, ligands which bind vascular endothelial growth factor (VEGF) receptors and show promise as potential molecular imaging agents and angiogenesis therapeutics. Affibody variants were selected from newly designed libraries based on the original monomeric molecules and an in silico approach was used in parallel to identify potentially stabilizing mutations. The selected mutants were characterized by CD spectroscopy and biosensor assays and the most promising ones combined into new monomeric constructs. CD data showed that different stabilizing mutations had an additive effect. Ultimately, dimeric fusion constructs were created with the capability to fully refold after heat-induced unfolding and which exhibited a remarkable increase in melting temperature by 15°C.
Güler, Svedmark, Abouzayed, Orlova, and Löfblom. 2020. “Increasing Thermal Stability and Improving Biodistribution of VEGFR2-Binding Affibody Molecules by a Combination of in Silico and Directed Evolution Approaches.” Scientific Reports 10: 18148.
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