Design of Lipid–Protein Conjugates Using Amphiphilic Peptide Substrates of Microbial Transglutaminase

Lipid modification of proteins plays a significant role in regulating the cellular environment. Mimicking natural lipidated proteins is a key technique for assessing the function of proteins modified with lipids and also to render self-assembly of lipids to a target protein. Herein, we report a facile method of conjugating proteins with lipid-fused peptides under homogeneous physiological conditions by using the microbial transglutaminase (MTG) reaction. MTG catalyzes the cross-linking reaction between a specific glutamine (Q) in a protein and a lysine (K) in newly designed lipid-fused peptides. The water-soluble peptide substrates for lipid modification, C₁₄-X-MRHKGS, were newly synthesized, where C₁₄, X, and MRHKGS represent myristic acid, linker peptides composed of G, P, or S, and MTG-reactive K surrounded with basic amino acids, respectively. The MTG-mediated cross-linking reaction between a protein fused with LLQG at the C-terminus and C₁₄-X-MRHKGS (5 molar eq) dissolved in a phosphate saline solution resulted in lipid–protein conjugates with yields of 70 to 100%. The anchoring ability of the obtained lipid–protein conjugates to cell membranes was dependent on the number of G residues in the G_<i>n</i>S linker, suggesting that self-assembly and hydrophobicity of the G_nS motif serves to enhance membrane anchoring of lipid–protein conjugates.