4 years ago

Proline-rich chaperones are compared computationally and experimentally for their abilities to facilitate recombinant butyrylcholinesterase tetramerization in CHO cells

Juechun Tang, Michael J. Betenbaugh, Cheng-yu Chung, Qiong Wang, Charles H. Chen, Jeffrey H. Chu, Martin B. Ulmschneider, Joseph Priola
Human butyrylcholinesterase (BChE), predominantly tetramers with a residence time of days, offers the potential to scavenge organophosphorus pesticides and chemical warfare agents. Efficient assembly of human BChE into tetramers requires an association with proline-rich peptide chaperones. In this study, the incorporation of different proline-rich peptide chaperones into BChE was investigated computationally and experimentally. First, we applied molecular dynamic (MD) simulations to interpret the interactions between proline-rich chaperones with human BChE tetramer domains. The P24 chaperone which contains 24 prolines, promoted the association of BChE tetramer with a 74 % simulated helicity of BChE subunits, whereas the control without chaperone and BChE with an 8-proline chaperone (P8) complex exhibited 55.8 % and 60.6 % predicted helicity, respectively. The interaction of proline-rich chaperones with BChE subunits (B-P) provides a conduit to facilitate the interactions between BChE subunits (B-B) of the complex, which is mainly attributed to hydrophobic interactions and hydrogen-bond binding. Experimental assessment of these two proline-rich chaperones plus a 14-proline chaperone (P14) was performed and confirmed that P24 has superior capability to facilitate recombinant BChE (rBChE) tetramerization with >60 % rBChE tetramer in P24-transfected rBChE cells, whereas P14- and P8-transfected rBChE cells have 44 % and 33 % rBChE tetramer respectively. The rBChE control had 14% tetramer. Finally, we developed a stable rBChE tetramer expression system in CHO cells by enriching P24 expression in rBChE expressing cells. Overall, our simulations provided a design concept for identifying proline-rich peptides that promote the rBChE tetramerization in CHO cells.

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/biot.201700479

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