Effect of heat treatment on protein for biosensing using glycopolymer surface

A B S T R A C T Glycopolymers with different glycan ratios of galactose and mannose were synthesized using the reversible addition–fragmentation chain-transfer (RAFT) polymerization and directly immobilized as biorecognition elements for the lectin peanut agglutinin (PNA). To investigate the influence of protein structural stability on binding affinity, PNA samples were heat-treated prior to analysis using surface plasmon resonance imaging (SPRi). The SPRi results obtained from glycopolymer-immobilized surfaces demonstrated that PNA binding was dependent on galactose density in the glycopolymer. Meanwhile, the association constant ( K a ) was influenced by the cluster-glycoside effect formed between the glycopolymer layer and the PNA. Heat treatment above 30 °C led to a decrease in binding affinity and an increase in nonspecific interactions due to structural destabilization and protein unfolding. These findings highlight the importance of protein structural integrity in lectin–glycopolymer interactions and provide valuable insight for the development of reliable glycopolymer-based biosensors.