Purification, characterization, and N-glycosylation of recombinant butyrylcholinesterase from transgenic rice cell suspension cultures.

Autor: Corbin JM; Department of Chemical Engineering, University of California, Davis, California., Kailemia MJ; Department of Chemistry, University of California, Davis, California., Cadieux CL; Medical Toxicology Division, US Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, Maryland., Alkanaimsh S; Department of Chemical Engineering, University of California, Davis, California.; Department of Chemical Engineering, College of Engineering and Petroleum, Kuwait University, Safat, Kuwait., Karuppanan K; Department of Chemical Engineering, University of California, Davis, California., Rodriguez RL; Department of Molecular and Cellular Biology, University of California, Davis, California.; Global HealthShare Initiative, University of California, Davis, California., Lebrilla CB; Department of Chemistry, University of California, Davis, California., Cerasoli DM; Medical Toxicology Division, US Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, Maryland., McDonald KA; Department of Chemical Engineering, University of California, Davis, California.; Global HealthShare Initiative, University of California, Davis, California., Nandi S; Department of Chemical Engineering, University of California, Davis, California.; Global HealthShare Initiative, University of California, Davis, California.
Jazyk: angličtina
Zdroj: Biotechnology and bioengineering [Biotechnol Bioeng] 2018 May; Vol. 115 (5), pp. 1301-1310. Date of Electronic Publication: 2018 Feb 27.
DOI: 10.1002/bit.26557
Abstrakt: Recombinant butyrylcholinesterase produced in a metabolically regulated transgenic rice cell culture (rrBChE) was purified to produce a highly pure (95%), active form of enzyme. The developed downstream process uses common manufacturing friendly operations including tangential flow filtration, anion-exchange chromatography, and affinity chromatography to obtain a process recovery of 42% active rrBChE. The purified rrBChE was then characterized to confirm its comparability to the native human form of the molecule (hBChE). The recombinant and native enzyme demonstrated comparable enzymatic behavior and had an identical amino acid sequence. However, rrBChE differs in that it contains plant-type complex N-glycans, including an α-1,3 linked core fucose, and a β-1,2 xylose, and lacking a terminal sialic acid. Despite this difference, rrBChE is demonstrated to be an effective stoichiometric bioscavenger for five different organophosphorous nerve agents in vitro. Together, the efficient downstream processing scheme and functionality of rrBChE confirm its promise as a cost-effective alternative to hBChE for prophylactic and therapeutic use.
(© 2018 Wiley Periodicals, Inc.)
Databáze: MEDLINE
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