Separation in a single step by affinity chromatography of cholinesterases differing in subunit number.
We describe an affinity chromatography method in which dimethylaminoethylbenzoic acid-Sepharose 4B is used, making it possible to separate in one step the molecular forms of globular acetylcholinesterase (AChE, EC 3.1.1.7) or butyrylcholinesterase (ChE, EC 3.1.1.8). A crude extract containing these enzymes was deposited onto the chromatography gel, washed, and eluted by a linear gradient of tetramethylammonium chloride (0-0.3 M). With rat brain AChE, two well-separated peaks were eluted in the presence of 1% Triton X-100; the first peak corresponded to 4 S forms and the second to 11 S forms. This separation was very efficient for salt-soluble activity and less efficient for the detergent-soluble AChE. In this case, the 4 S peak represented only 6.5% of total detergent-soluble activity and was cross-contaminated by the 11 S form. Rat serum ChE was efficiently separated into two peaks of 7 S and 11 S. This method could potentially be adapted to separate other multimeric proteins with varying numbers of affinity sites.
Andres
C
C
Centre de Neurochimie, CNRS, Strasbourg, France.
el Mourabit
M
M
Mark
J
J
Waksman
A
A
D000110:Acetylcholinesterase / Q000737:chemistry / Q000302:isolation & purification; D000818:Animals; D001921:Brain / Q000201:enzymology; D002091:Butyrylcholinesterase / Q000097:blood / Q000737:chemistry; D002802:Cholinesterases / Q000737:chemistry / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods*; D011487:Protein Conformation; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility
D000110:Acetylcholinesterase; D002091:Butyrylcholinesterase; D002802:Cholinesterases