Hupresin Retains Binding Capacity for Butyrylcholinesterase and Acetylcholinesterase after Sanitation with Sodium Hydroxide

Abstract

Hupresin is a new affinity resin that binds butyrylcholinesterase (BChE) in human plasma and acetylcholinesterase (AChE) solubilized from red blood cells (RBC). Hupresin is available from the CHEMFORASE company. BChE in human plasma binds to Hupresin and is released with 0.1 M trimethylammonium bromide (TMA) with full activity and 10–15% purity. BChE immunopurified from plasma by binding to immobilized monoclonal beads has fewer contaminating proteins than the one-step Hupresin-purified BChE. However, when affinity chromatography on Hupresin follows ion exchange chromatography at pH 4.5, BChE is 99% pure. The membrane bound AChE, solubilized from human RBC with 0.6% Triton X-100, binds to Hupresin and remains bound during washing with sodium chloride. Human AChE is not released in significant quantities with non-denaturing solvents, but is recovered in 1% trifluoroacetic acid. The denatured, partially purified AChE is useful for detecting exposure to nerve agents by mass spectrometry. Our goal was to determine whether Hupresin retains binding capacity for BChE and AChE after Hupresin is washed with 0.1 M NaOH. A 2 mL column of Hupresin equilibrated in 20 mM TrisCl pH 7.5 was used in seven consecutive trials to measure binding and recovery of BChE from 100 mL human plasma. Between each trial the Hupresin was washed with 10 column volumes of 0.1 M sodium hydroxide. A similar trial was conducted with red blood cell AChE in 0.6% Triton X-100. It was found that the binding capacity for BChE and AChE was unaffected by washing Hupresin with 0.1 M sodium hydroxide. Hupresin could be washed with sodium hydroxide at least seven times without losing binding capacity.

Effects of Kifunensine on Production and <em>N</em>-glycosylation Modification of Butyrylcholinesterase in a Transgenic Rice Cell Culture Bioreactor

“…It could be implied (Figure 3a, lane CA-) through image analysis, which is consistent with our previous reports using the same steps [2,3]. However, culture medium rrBChE in the absence of kifunensine was only 31% purity (Figure 3a with a single step Hupresin ® , while 99% purity of hBChE was achieved when Q-ceramic ion exchange chromatography at pH 4.5 was used prior the Hupresin ® step [42]. Thus, to acquire high rrBChE purity in the Hupresin ® elution, it may be advantageous to enrich rrBChE purity in the Hupresin ® loading solution.…”

Effects of Kifunensine on Production and N-Glycosylation Modification of Butyrylcholinesterase in a Transgenic Rice Cell Culture Bioreactor

“…Alkanaimsh et al also found contaminating protein bands from Hupresin ® elution due to nonspecific binding when nonenriched Nicotiana benthamiana rBChE cell extract was loaded onto Hupresin ® [ 2 ]. Starting with low BChE purity, Onder et al obtained 10–15% purity of hBChE purified from human blood plasma with a single-step Hupresin ® , while 99% purity of hBChE was achieved when Q-ceramic ion exchange chromatography at pH 4.5 was used prior to the Hupresin ® step [ 48 ]. Thus, to acquire high rrBChE purity in the Hupresin ® elution, it may be advantageous to enrich rrBChE purity in the Hupresin ® loading solution.…”

Purification of human butyrylcholinesterase from frozen Cohn fraction IV-4 by ion exchange and Hupresin affinity chromatography

“…A small scale trial tested the effect of sanitation with 0.1 M sodium hydroxide on the binding of BChE from 100 mL of human plasma by 2 mL Hupresin. We reported that 7 rounds of sanitation had no adverse effect [17].…”