Table 2. Calculated Michaelis-Menten parameters for chlorzoxazone 6-hydroxylation by wild-type and variant forms of CYP2E1.

Each value is the mean ± SD of three independent experiments. Vmax represents the maximal rate at which the enzyme metabolizes the substrate; Km represents the Michaelis constant; and the Clint value represents the intrinsic clearance. Three online software programs were simultaneously used to predict genetic mutations. The three algorithms predicted no effect (benign/tolerant/neutral) or loss of function (damaging/probably damaging/deleterious). When the results of two of the three software predictions were consistent with the results of the HPLC experiment, the results were considered to be consistent, and we added YES; otherwise, we added NO.

Variant	Vmax± SD (pmol/min/pmol CYP2E1)	Km± SD (µM)	CLint (µL/min/pmol CYP2E1) (% of wild-type)	Polyphen	SIFT	PROVEAN	Concordance Y/N
Wild-type	21.75 ± 1.413	63.47 ± 13.81	0.3841 ± 0.0631
c.1263C>T	21.65 ± 1.393	56.95 ± 12.67	0.4027 ± 0.052 (102%)	N/A	Tolerated	Neutral	YES
c.1009C>T	0.02			N/A	N/A	Deleterious	N/A
c.[517G>A+1263C>T]	21.34 ± 1.29	60.81 ± 12.46	0.379 ± 0.078 (98.6%)	N/A	N/A	N/A	N/A
c.517G>A	23.06 ± 1.354	53.2 ± 11.03*	0.511 ± 0.089 (131%)**	probably damaging	Tolerated	Deleterious	YES
c.[227G>A+1263C>T]	14.54 ± 0.8015**	64.09 ± 11.82	0.258 ± 0.055 (67.2%)**	N/A	N/A	N/A	N/A
c.227G>A	12.96 ± 0.74**	65.8 ± 12.45	0.22 ± 0.041 (57.2%)**	probably damaging	Damaging	Neutral	YES

Notes.

^*P ≤ 0.05.

^**P ≤ 0.01.

^***P ≤ 0.001.