Purification of recombinant human butyrylcholinesterase on Hupresin®.

ABSTRACT: Affinity chromatography on procainamide-Sepharose has been an important step in the purification of butyrylcholinesterase (BChE) and acetylcholinesterase (AChE) since its introduction in 1978. The procainamide affinity gel has limitations. In the present report a new affinity gel called Hupresin® was evaluated for its ability to purify truncated, recombinant human butyrylcholinesterase (rHuBChE) expressed in a stably transfected Chinese Hamster Ovary cell line. We present a detailed example of the purification of rHuBChE secreted into 3940?mL of serum-free culture medium. The starting material contained 13,163?units of BChE activity (20.9?mg). rHuBChE was purified to homogeneity in a single step by passage over 82?mL of Hupresin® eluted with 0.1?M tetramethylammonium bromide in 20?mM TrisCl pH?7.5. The fraction with the highest specific activity of 630?units/mg contained 11?mg of BChE. Hupresin® is superior to procainamide-Sepharose for purification of BChE, but is not suitable for purifying native AChE because Hupresin® binds AChE so tightly that AChE is not released with buffers, but is desorbed with denaturing solvents such as 50% acetonitrile or 1% trifluoroacetic acid. Procainamide-Sepharose will continue to be useful for purification of AChE.

SUBMITTER: Lockridge O 

PROVIDER: S-EPMC6275104 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature