Abstract
One enzyme family, the transglutaminase could potentially be involved in the covalent cross-linking of the substrate proteins on biological membranes. The ε-(γ-glutamyl)lysine bonds between Gln and Lys of proteins, formed by transglutaminase, are entirely resistant to chemical and physical disruption. The cross-linking reaction of various substrate proteins by the transglutaminase was investigated in order to establish the method to control the cross-linking reaction of the substrate proteins on the model membrane (liposomes) under the environmental stress conditions. The cross-linking reaction of various substrate proteins at various conformational states was firstly analyzed by using SDS-PAGE, resulting that the rate constant of the cross-linking reaction was highly dependent on the conformational state of the proteins under the given stress conditions. The increase of the local hydrophobicity of the proteins, which is corresponding with the structural fluctuation, is found to be one of the possible factors, which can be analyzed by aqueous two-phase partitioning method. Based on the protein structure, it is suggesting that the rate constant of the cross-linking reaction was furthermore dependent on the hydrophobicity of the Gln residues of the substrate proteins to be cross-linked. The effect of the addition of various liposome was also investigated, implying that the fluidity of liposome may also affect the reaction kinetics. Based on these results, a strategy to control the cross-linking of the substrate protein was finally been discussed.