Nanogel artificial chaperone: Refolding and cell-free protein synthesis of GFP

POLY 421

Wakiko Asayama1, Yuta Nomura1, Shin-ichi Sawada1, Nobuyuki Morimoto1, and Kazunari Akiyoshi, akiyoshi.org@tmd.ac.jp2. (1) Department of Organic Materials, Institute of Biomaterials & Bioengineering, Tokyo Medical & Dental University, 2-3-10, Kanda-surugadai, Chiyoda-ku, Tokyo, 101-0062, Japan, (2) Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, 2-3-10, Kanda-surugadai, Chiyoda-ku, Tokyo, 101-0062, Japan
In biotechnology, there are many problems in protein folding. In living system, molecular chaperones selectively trap denatured proteins or their intermediates to prevent their irreversible aggregation. To simulate the function of molecular chaperones, we developed a novel artificial chaperone using nanogels. Nanogels of cholesterol-bearing pullulan (CHP) complexed with proteins to effectively prevented protein aggregation, and released proteins by addition of cyclodextrins. We report here the effect of CHP nanogels as artificial chaperone for the refolding of acid-denatured GFP and for E.coli cell-free protein synthesis. The chaperon-like activity of nanogels is comparable to that of GroEL in the refolding of acid-denatured GFP. Furthermore, CHP nanogel trapped nascent GFP in E.coli cell-free protein synthesis system and released native GFP after the addition of methyl-β-CD without influence to the GFP synthesis system. The nanogel system is a useful tool in high expression methods for easily aggregated and unstable proteins.
 

7th International Biorelated Polymers Symposium
6:00 PM-8:00 PM, Tuesday, 12 September 2006 Moscone Center -- Hall D, Poster

Division of Polymer Chemistry

The 232nd ACS National Meeting, San Francisco, CA, September 10-14, 2006