TABLE 1.

Kinetic parameters for the butyl-FLIP activity of B. cereus PI-PLC obtained by fitting to the two-site model

Data set	Curve shape	No. of fitted parameters	diC6PC (mM)	KS (mM)	α′	(mM)	kp (s−1)	β′	KA (mM)	α	β
Fig. 4 A Solid line	Sigmoidal	5	0	0.5	0.2	0.15	240	1.4	—	—	—
Fig. 4 B Dotted line	Hyperbolic (approx.)	3	1.07	0.5	0.2	0.15	218	1.4	0.2	0.03	1.0
Fig. 4 B Solid line	Hyperbolic (approx.)	7	1.07	0.5	0.1	0.04	220	1.5	0.2	0.02	0.9
Fig. 5	Hyperbolic	7	0–1	0.5	0.2	0.15	100	1.4	0.2	0.02	1.0

The data of Fig. 4 were simulated with a five-parameter (K_S, α′, β′, k_p) fit using Eq. 1, or equivalently using Eq. 5 with [A] = [diC₆PC] = 0. K_A was held constant at 0.2 mM. Fig. 4 B was simulated with Eq. 5 by two methods. In the first method, the four parameters (K_S, α′, β′) were fixed at values arrived at for Fig. 4 A, and only three parameters (k_p, α, β) were allowed to vary. In the second method, all seven parameters (K_S, α′, β′, k_p, α, β) were allowed to vary. The fit to the data of Fig. 5 was carried out with Eq. 5, varying the same seven parameters, and [S] = 8.6 μM butyl-FLIP. Values were arrived at by visual curve fitting and are thus semiquantitative. Estimated errors are 10% for K_S and 20% for α′, β′, α, β. The variation in k_p was larger due to differences in enzyme activity from different preparations (see text).