Development of Polymerized Liposomes for Protein Recognition

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INTELLECTUAL MERIT: It is proposed to develop liposomes that have been designed to recognize specific proteins, the surface topology of which has been imprinted on the surface of the liposomes. The targeted protein for this study is matrix metalloproteinase-9, which is involved in a number of physiological and pathological processes. Liposomes will be constructed using a variety of lipids, cationic, neutral, and anionic, in order to permit tuning of the liposome so that its gel transition temperature lies above the temperature at which the target protein undergoes thermal denaturation. Lipids possessing functionality suitable for photopolymerization will be included in the lipid mixture. Some of the lipids will also have head groups that bind specifically to the enzyme so as to orient the protein uniquely with respect to the liposome surface. During incubation of the complex at a temperature intermediate between the gel transition of the liposome and the denaturation temperature of the protein, the fluidity of the membrane will permit the liposome to accommodate its surface to the surface of the protein and form a complementary template for later binding of that protein. Cooling below the gel transition of the liposome and photopolymerization of the liposome membrane then locks in the template. Preliminary work shows that the affinity of the enzyme for the templated liposome is enhanced by one - two orders of magnitude relative to the untemplated liposome. The work will proceed as follows: (i) synthesize structurally diverse, polymerizable lipids to formulate liposomes for isozyme-selective detection and inhibition of matrix metalloproteinase-9 (MMP-9), (ii) formulate polymerized liposomes with different types of head groups to serve as highly specific multi-prong inhibitors for MMP isozymes, (iii) develop fluorescence based sensors using chelated Eu(III) and Tb(III) for the isozyme-selective detection of MMP-9, (iv) determine the mechanisms of the liposome assisted detection and inhibition of MMP-9BROADER IMPACTS: The proposed work opens a new avenue to molecular imprinting, especially in the area of protein recognition and "artificial antibody" preparation. Applications can be imagined in liposome-based diagnostic tools for biological warfare agents, detection of plant and animal pathogens, and development of prophylactic and therapeutic agents. The PIs will support two Native American undergraduates working in the lab. Both will host high school students and public school teachers in their laboratories in NDSU-sponsored summer research programs.