Peroxo-iron and Oxenoid-iron Species as Alternative Oxygenating Agents in Cytochrome P450-catalyzed Reactions: Switching by Threonine-302 to Alanine Mutagenesis of Cytochrome P450 2B4.
- UNCG Author/Contributor (non-UNCG co-authors, if there are any, appear on document)
- Gregory M. Raner, Associate Professor and Graduate Director (Creator)
- Institution
- The University of North Carolina at Greensboro (UNCG )
- Web Site: http://library.uncg.edu/
Abstract: ABSTRACT Among biological catalysts, cytochrome
P450 is unmatchedi n its multiplicityo f isoforms,i nducers,
substrates,a nd typeso f chemical reactionsc atalyzed.I n the
presents tudy,e videncei s givent hatt hisv ersatilityex tendst o
the nature of the active oxidant. Althoughm echanistice vidence
froms everal laboratoriesp ointst o a hypervalentir onoxenoid
species in P450-catalyzed oxygenation reactions,
Akhtar and colleagues [Akhtar,M ., Calder, M. R., Corina,
D. L. & WrightJ, . N. (1982) BiochemJ. . 201,5 69-580] proposed
that in steroidd eformylatioenf fectedb y P450 aromatase an
iron-peroxos pecies is involvedW. e have shownm orer ecently
that purifiedl iver microsomalP 450 cytochromesi,n cluding
phenobarbital-inducedP 450 2B4, catalyze the analogous deformylationo
f a series of xenobiotica ldehydes with olefin
formationT. he investigationp resentedh ere on the effecto f
site-directedm utagenesiso f threonine-302t o alanine on the
activitieso f recombinantP 450 2B4 with N-terminala mino
acids 2-27 deleted [2B4 (A2-27)] makes use of evidencef rom
otherl aboratoriest hat the correspondingm utationi n bacterial
P450s interferesw ith the activationo f dioxygent o the
oxenoid species by blockingp rotond eliveryt o the active site.
The rates of NADPH oxidation,h ydrogenp eroxide production,
a nd productf ormationf romf ours ubstrates,i ncluding
formaldehydef rom benzphetamineN -demethylationa,c etophenone
from1 -phenylethanoolx idation,c yclohexanolf rom
cyclohexaneh ydroxylationan, d cyclohexenef romc yclohexane
carboxaldehyded eformylationw, ere determinedw ith P450s
2B4, 2B4 (A2-27), and 2B4 (A2-27) T302A. Replacement of
the threoniner esidue in the truncatedc ytochromge ave a 1.6-
to 2.5-foldi ncrease in peroxidef ormationin the presenceo fa
substrate,b ut resultedi n decreased productf ormationf rom
benzphetamine (9-fold), cyclohexane (4-fold), and 1-phenylethanol
(2-fold).I n sharp contrast,t hed eformylatioonf c yclohexane
carboxaldehydeb y the T302A mutantw as increased
about 10-foldO. n the basis oft hesef indingsa nd our previous
evidencet hat aldehyded eformylationis supportedb y added
H202, but not by artificial oxidants, we conclude that the
iron-peroxys pecies is the directo xygend onor. It remains to
be established which of the many other oxidative reactions
involvingP 450 utilize this species and the extentt o which
peroxo-irona nd oxenoid-ironf unctiona s alternativeo xygenating
agents with the numerous isoforms of this versatile
catalyst.
Peroxo-iron and Oxenoid-iron Species as Alternative Oxygenating Agents in Cytochrome P450-catalyzed Reactions: Switching by Threonine-302 to Alanine Mutagenesis of Cytochrome P450 2B4.
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Additional Information
- Publication
- Language: English
- Date: 1996
- Keywords
- biological catalysts, peroxo-iron, oxenoid-iron, cytochrome P450, biochemistry