Table 5

Kinetic analysis of the 4- and 2-hydroxylation of 17β-estradiol by human CYP1B1 variants expressed in S. cerevisiae

CYP1B1 variant	Amino acid substitutions	Expression level	K_m		V_max		V_max/K_mratio
CYP1B1 variant	Amino acid substitutions	Expression level	4-Hydroxy	2-Hydroxy	4-Hydroxy	2-Hydroxy	4-Hydroxy	2-Hydroxy
		pmol/mg	μM		pmol/pmol P450/min
CYP1B1.1	None	14.6	5.3 ± 0.3	8.9 ± 1.1	3.8 ± 0.1	0.5 ± 0.10	0.71	0.05
CYP1B1.2	Arg48Gly, Ala119Ser	3.6	6.4 ± 1.9	7.7 ± 0.1	4.7 ± 0.2	0.5 ± 0.10	0.73	0.06
CYP1B1.3	Leu432Val;	7.1	6.4 ± 0.2	8.7 ± 2.4	3.3 ± 0.2	0.65 ± 0.10	0.51	0.07
CYP1B1.4	Asn453Ser	5.2	7.0 ± 1.0	9.4 ± 1.8	3.6 ± 0.6	0.55 ± 0.06	0.51	0.06
CYP1B1.6	Arg48Gly, Ala119Ser, Leu432Val	16.1	12.1 ± 1.0^5-b	16.6 ± 0.1^5-a	2.1 ± 0.1^5-b	0.3 ± 0.05	0.17^5-c	0.01^5-a
CYP1B1.7	Arg48Gly, Ala119Ser, Leu432Val, Ala443Gly	7.4	13.4 ± 1.3^5-b	39.1 ± 2.4^5-b	1.7 ± 0.2^5-b	0.7 ± 0.08	0.12^5-c	0.01^5-b
L432V ⁺ N453S	Leu432Val, Asn453Ser	11.5	7.9 ± 0.6	7.5 ± 0.5	2.6 ± 0.1	0.3 ± 0.05	0.32	0.04
L432V ⁺ A443G	Leu432Val, Ala443Gly	10.0	8.9 ± 0.6	9.7 ± 1.7	3.1 ± 0.1	0.5 ± 0.10	0.34	0.05

Kinetic parameters are presented as means ± S.D. from three independent analyses using two separate microsomal preparations for each enzyme variant.

↵5-a Significant difference (p < 0.05) compared with CYP1B1.1 using unpairedt test.
↵5-b Significant difference (p < 0.01) compared with CYP1B1.1 using unpairedt test.
↵5-c Significant difference (p < 0.001) compared with CYP1B1.1 using unpairedt test.