Development of Multi-functional Protein Nanostructures for Biomedical Applications

Abstract

Department of Biological SciencesRecent development of construction of nanostructured materials such as nanoparticles, nanowire, and nanosheets has made a great contribution to the advance of our life since they are closely related to the various biomedical applications. Despite the recent advances in technology for constructing complex nanostructured materials, there is still a long way to go to develop more advanced and more sophisticated materials. Construction of nano-sized supramolecules with precise orientation of structures and functions can provide opportunity to develop new materials which can control the complex biological processes. In this study, we used protein cage nanoparticle as the template for the construction of the complex nanostructured materials and utilized them in various biomedical fields. The Aquifex aeolicus lumazine synthase protein cage nanoparticle was engineered to enable post-translational surface modification with various proteins. SpyTag (ST) displayed lumazine synthase forms covalent display of various SpyCatcher (SC) fusion proteins, and those protein cage nanoparticles were utilized as building blocks for the construction of enzyme-containing multi-layered 3D nanoreaction clusters with enhanced enzymatic activity. The Thermotoga maritima encapsulin protein cage nanoparticle, having outer diameter of 24 nm and inner diameter of 20 nm, was engineered for the simultaneous modification of interior space and exterior surface. Introduction of both split intein fragments and ST/SC enabled protein cargo encapsulation and additional ligand decoration, respectively, in a mix-and-match manner. The constructed protein nanostructures were further applied in the enzyme immobilization, multi-layer construction, and targeted cell imaging. Furthermore, previously developed ST displayed lumazine synthase protein cage nanoparticle was utilized as a template for simultaneous immobilization of the potential therapeutic enzyme, lactate oxidase (LOX) and catalase (CAT), to modulate the tumor microenvironment for the enhanced tumor therapy. The constructed multi-enzyme complex effectively consumed tumor lactate even in the hypoxic conditions, showing the potential usage in tumor treatment which can induce the reprogramming the tumor microenvironment and activation of immune responses against tumors. The approaches we described here may provide new opportunities to construct protein cage nanoparticle-based complex nanostructured materials utilized for various biomedical applications and nanostructured biosensor devicesclos