Abstract
Human liver microsomes and recombinant human P450 have been used as enzyme source in order to better understand the requirement for the optimal rate of omega and (omega;-1)-hydroxylations of fatty acids by cytochromes P450 2E1 and 4A. Three parameters were studied: alkyl chain length, presence and configuration of double bond(s) in the alkyl chain, and involvement of carboxylic function in the fatty acid binding inside the access channel of P450 active site. The total rate of metabolite formation decreased when increasing the alkyl chain length of saturated fatty acids (from C12 to C16), while no hydroxylated metabolite was detected when liver microsomes were incubated with stearic acid. However, unsaturated fatty acids, such as oleic, elaidic and linoleic acids, were omega and (omega;-1)-hydroxylated with an efficiency at least similar to palmitic acid. The (omega;-1)/omega ratio decreased from 2.8 to 1 with lauric, myristic and palmitic acids as substrates, while the reverse was observed for unsaturated C18 fatty acids which are mainly omega-hydroxylated, except for elaidic acid showing a metabolic profile quite similar to those of saturated fatty acids. The double bond configuration did not significantly modify the ability of hydroxylation of fatty acid, while the negatively charged carboxylic group allowed a configuration energetically favourable for omega and (omega;-1)-hydroxylation inside the access channel of active site.
Highlights
Human liver microsomes and recombinant human cytochrome P450 or CYP (P450) have been used as enzyme source in order to better understand the requirement for the optimal rate of and (–1)-hydroxylations of fatty acids by cytochromes P450 2E1 and 4A
The RP-HPLC profiles of fatty acids incubated with human liver microsomes showed the presence of two major metabolites with retention times corresponding to (–1) and -hydroxylated metabolites
Two P450 isoforms have been investigated in the hydroxylations of fatty acids, namely CYP2E1 and CYP4A that are shown to be highly regioselective for catalyzing (–1) and -hydroxylations, respectively
Summary
Human liver microsomes and recombinant human P450 have been used as enzyme source in order to better understand the requirement for the optimal rate of and (–1)-hydroxylations of fatty acids by cytochromes P450 2E1 and 4A. Three parameters were studied: alkyl chain length, presence and configuration of double bond(s) in the alkyl chain, and involvement of carboxylic function in the fatty acid binding inside the access channel of P450 active site. The (–1)/ ratio decreased from 2.8 to 1 with lauric, myristic and palmitic acids as substrates, while the reverse was observed for unsaturated C18 fatty acids which are mainly -hydroxylated, except for elaidic acid showing a metabolic profile quite similar to those of saturated fatty acids. The double bond configuration did not significantly modify the ability of hydroxylation of fatty acid, while the negatively charged carboxylic group allowed a configuration energetically favourable for and (–1)-hydroxylation inside the access channel of active site.—Adas, F., J.P. Salaün, F.
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