Figure 4

CYP110 Absorbance Spectra, Titrated Spectral Shift, and Double Reciprocal Plot in the presence and absence of various hydrocarbons. The absorbance characteristics were monitored for the purified soluble CYP110 from E. coli. The sample was diluted 1:5 in buffer A with 0.2 % deoxycholate. To reduce the P450, 5 mg of sodium dithionate was added to the sample. Carbon monoxide was then bubbled into the sample for 30 seconds. Panel A. Oxidized, reduced and carbon monoxide bound spectra. Panel B. Carbon monoxide difference spectrum with a peak at 450 nm. Panel C. The oxidized spectra for 3 μM CYP110 in buffer A + 0.2 % deoxycholate and titrated with tetradecanoic acid. The concentrations of tetradecanoic acid shown are 0, 0.1, 0.2, 0.5, 0.7, and 1.0 mM. Panel D. (Top) The difference spectra generated for CYP110 when titrated with tetradecanoic acid. The bottom plot for Panel D shows the spectrum without fatty acid subtracted from each spectra with fatty acid. The double reciprical plot of 1/ΔAbs vs 1/S is shown and the X-intercept is calculated by linear regression. Ks is defined as -1/X-intercept. ΔAbs for each fatty acid concentration is defined as the resulting value at 388 nm.