Butyrl Cholinesterase catalyzes the hydrolysis of a number of choline esters according to the following reaction:
Acylcholine + H2O → choline + acid
It is a tetrameric glycoprotein with four subunits of equal size, each having a molecular weight of 110 kDa. Butyryl cholinesterase hydrolyzes butyrylcholine four times more rapidly than acetylcholine. Unlike acetyl cholinesterase, it does not hydrolyze D-≥-methyl acetylcholine. It is inhibited by 10-5 M physostigmine, numerous organophosphate esters, the carbamate derivates and quaternary ammonium salts.
Specificity
The enzyme is more active with butyryl and propionyl choline than with acetyl choline. Non-choline esters, such as procaine, morphine esters, atropine and cocaine, are susceptible to the action of ButChE.
Composition
ButChE presents a tetrameric structure with equally sized subunits of 110,000 daltons. It is a glycoprotein. The amino acid and carbohydrate composition has been determined based upon a particle weight of 101,000.
Characteristics
Molecular Weight: 440,000
Optimal pH: 6.0–8.0
Extinction Coefficient: E280 = 13.6
Activators: Ca²⁺ and Mg²⁺
Inhibitors: Numerous organophosphate esters, carbamate derivatives and quaternary ammonium salts.
Assay
Method
The reaction velocity is determined by a titrimetric measurement of the acid released during the hydrolysis of acetylcholine. One Unit causes hydrolysis of one micromole of acetylcholine per minute at 25°C and pH 7.4, under the specified conditions.
Reagents
0.02 M Tris⋅HCl buffer, pH 7.4
0.2 M Magnesium chloride
2.2 M Acetylcholine chloride
Titrant: 0.01-0.02 N NaOH, standardized
Enzyme
Dissolve at one mg/ml in reagent grade water.
Procedure
The titration can be measured with either an automatic titrator or with a laboratory pH meter. The reaction vessel should be maintained at 25°C.
Pipette the following into a titration vessel at 25°C
Reagent grade water 7.0 ml
0.2 M Magnesium chloride 3.0 ml
0.02 M Tris⋅HCl buffer, pH 7.4 3.0 ml
Enzyme 0.1-0.5 ml
Adjust pH to 7.5 with titrant. At zero time add 1 ml of 2.2 M acetylcholine and adjust pH to 7.4. Record the amount of standard base required to maintain the pH at 7.4 for 5-6 minutes after a constant rate is achieved.
Calculation
J2622133
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