. Author manuscript; available in PMC: 2008 Apr 15.

Published in final edited form as: Arch Biochem Biophys. 2006 Dec 3;460(2):262–273. doi: 10.1016/j.abb.2006.11.018

Table III.

Substrate binding properties of recombinant rat CY24A1 wild-type and serine-57 mutants for vitamin D3 metabolites and 3-OH analogs.

Substrate	Enzyme	Spectral Perturbation^* (ΔAbs_{Max −}ΔAbs_Free)	Dissociation Constant Kd (μM)
25-(OH)D₃	WT	0.025 ± 0.002	0.311 ± 0.014
	S57D	0.029 ± 0.001	0.201 ± 0.008^e
	S57A	0.017 ± 0.003^e	0.320 ± 0.025

1,25-(OH)₂D₃	WT	0.044 ± 0.003	0.049 ± 0.006
	S57D	0.048 ± 0.002	0.013 ± 0.002^e
	S57A	0.031 ± 0.003^e	0.130 ± 0.008^e

3-epi-1,25-(OH)₂D₃	WT	0.011 ± 0.001	0.460 ± 0.013
	S57D	0.015 ± 0.001	0.421 ± 0.029
	S57A	0.012 ± 0.002	0.394 ± 0.024^d

3-deoxy-1,25-(OH)₂D₃	WT	0.032 ± 0.002	0.145 ± 0.008
	S57D	0.029 ± 0.001	0.136 ± 0.028
	S57A	0.039 ± 0.003^d	0.315 ± 0.016^e

ΔAbs_Free represents the initial difference in optical absorbance between 415 nm and 390 nm for the free enzyme prior to each titration.

ΔAbs_Max represents the maximal change in optical absorbance between 415 nm and 390 nm during each titration.

Mean ± SD for wild-type (WT) experiments (N=4 or greater). Values for mutants (N=3 or greater).

p < 0.05, compared to corresponding WT mean.

p < 0.001, compared to corresponding WT mean.

Measure of binding efficiency in the substrate pocket as reflected by perturbation of the heme spectra in the enzyme's active site.