An endogenous tetrameric wild-type human BChE expressed in Expi293 cells hydrolyzes the neutral substrate N-methylindoxyl acetate (NMIA) with the same K_m as wild-type huBChE (0.14 mM) [1]. For this enzyme, the steady state is preceded by a pre-steady state phase of several minutes in 10 mM Bis-Tris, pH 7 at 25°C.

Thermal inactivation of this BChE is biphasic. Kinetic constants (k₁ and k₂) for thermal inactivation shows differences between this mutant and plasma derived wtBChE: the Expi293 is more stable at 55°C and less stable at 60°C than natural wtBChE [2]. At 55°C half-life times of the first and the second phases are 11 min and 43 min for plasma wtBChE; 14 min and 36 min for the Expi293 wtBChE, respectively. At 60°C, the corresponding values are 6 min and 14 min for natural wtBChE; 3 min and 11 min for Expi293 wtBChE. The endogenous enzyme is more stable in urea: urea-induced denaturation is 10 % slower than for the wtBChE and the urea concentration at the mid-point of denaturation is 4.1M for wtBChE and 4.6M for the endogenous enzyme.

A bistable dynamic behavior of the endogenous BChE is also observed from pre-steady state behavior for hydrolysis of 1 mM NMIA, showing either long lags or bursts under the same conditions while plasma BChE shows only lags. Molecular mechanic simulations have been undertaken to determine the molecular basis of bistability of this wild-typeBChE.