title: Bio-inspired liposomal thrombomodulin conjugate through bio-orthogonal chemistry.
abstract: We report the synthesis of bioinspired liposomal thrombomodulin (TM) conjugates by chemoselective and site-specific liposomal conjugation of recombinant TM at C-terminus. TM is an endothelial cell membrane protein that acts as a major cofactor in the protein C anticoagulant pathway. To closely mimic membrane protein structural features of TM, we proposed membrane-mimetic re-expression of recombinant TM onto liposome. A recombinant TM containing the EGF-like 456 domains and an azidohomoalanine at C-terminus was expressed in E. coli. Conjugation of the recombinant TM onto liposome via Staudinger ligation and copper-free click chemistry were investigated as an optimal platform for exploring membrane protein TM's activity, respectively. The bioinspired liposomal TM conjugates were confirmed with Western blotting and protein C activation activity. The recombinant TM-liposome conjugates showed a 2-fold higher k(cat)/K(m) value for protein C activation than that of the recombinant TM alone, which indicated that the lipid membrane has a beneficiary effect on the recombinant TM's activity. The reported liposomal protein conjugate approach provides a rational design strategy for both studying membrane protein TM's functions and generating a membrane protein TM-based anticoagulant agent.
authors: Department of Chemistry, Cleveland State University, Cleveland, OH 44115, USA.
authors: Hailong
authors: H
authors: Zhang
authors: Jacob
authors: J
authors: Weingart
authors: Rui
authors: R
authors: Jiang
authors: Jianhao
authors: J
authors: Peng
authors: Qingyu
authors: Q
authors: Wu
authors: Xue-Long
authors: XL
authors: Sun
mesh_terms: D040761:Biomimetic Materials / Q000737:chemistry*; D032701:Biomimetics; D005456:Fluorescent Dyes / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry*; D015394:Molecular Structure; D009154:Mutation; D011994:Recombinant Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics; D018180:Thrombomodulin / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
chemical_list: D005456:Fluorescent Dyes; D008081:Liposomes; D011994:Recombinant Proteins; D018180:Thrombomodulin