0 citations0 references

Structure and Chemistry of Cytochrome P450

Chemical Reviews2005Vol. 105(6), pp. 2253–2278

Citations Over TimeTop 1% of 2005 papers

Ilia G. Denisov, Thomas M. Makris, Stephen G. Sligar, Ilme Schlichting

Abstract

The title to a seminar presentation by I. C. Gunsalus in 1973 was Oxygen: An essential toxin, referring to the complex \nrole that atmospheric dioxygen has in biology. The relatively simple function as terminal oxidant for aerobic life was dramatically \naugmented by Osamu Hayaishi with his identification of an enzyme that catalyzes the conversion of catechol to muconic acid by \noxidative cleavage.1 He named this biological catalyst pyrocatechase, which proved to be the landmark discovery of an enzyme \nthat incorporated atmospheric dioxygen into the carbon chain of the substrate, thereby initiating cleavage of the benzene ring. \nThis review of the oxygenase cytochrome P450 is dedicated to Dr. Hayaishi and his pioneering discovery in what is now the 50th anniversary of his work!\n\nWe now realize that Nature has found many ways to utilize atmospheric dioxygen to functionalize molecules through the use of a diverse \nset of cofactors. Flavin, non−heme iron, copper, and metalloporphyrin complexes have all been conscripted to metabolize atmospheric \ndioxygen in an oxygenase catalytic cycle, resulting in the incorporation of one or both oxygen atoms into a substrate. This review focuses \non one of the heme−containing classes, termed cytochrome P450s and abbreviated CYP. Although but one member in the large group of \noxygenases, the cytochrome P450s play a variety of critical roles in biology.\n\nMany members of the cytochrome P450 superfamily of hemoproteins are currently known, and the numbers continue to grow as more genomes \nare sequenced. There are almost 4000 identified P450 genes at the date of this writing, and they are collected and annotated in a variety \nof web sites, such as that maintained by Nelson (http://drnelson. utmem.edu/CytochromeP450.html). The cytochrome P450s have been found in \nall branches of the tree of life that catalogs the diversity of life forms. In the broadest terms, there are two main functional roles for \nthese oxygenases. One is the metabolism of xenobiotics (compounds exogenous to the organism) as a protective role of degradation or provision \nof polar handles for solubilization in preparation for excretion. A second broad functional role is in the biosynthesis of critical signaling \nmolecules used for control of development and homeostasis. In mammalian tissues the P450s play these roles through the metabolism of drugs \nand xenobiotics and the synthesis of steroid hormones and fat−soluble vitamin metabolism and the conversion of polyunsaturated fatty \nacids to biologically active molecules, respectively. Similar roles are fulfilled in plants (hormone biosynthesis and herbicide degradation) \nand insects (control of development via hormone biosynthesis or provision of insecticide resistance). For instance, plants have an unusually \nlarge number of P450 genes. A reason is their sessile nature: for example, plants defend themselves through breakdown of herbicides by \ncatalyzing the synthesis of a large number of secondary metabolites or by synthesizing defense molecules such as DIMBOA.2,3 In addition, \nthe biosynthesis of critical metabolic regulators is also often carried out by the cytochrome P450s.\n\nThe important metabolic role together with the unique chemistry and physical properties of the cytochrome P450s provide a strong attraction \nfor scientists in many disciplines. Relevance to human health was the initial focus of pharmacologists and toxicologists. The role of metal \ncenters and their associated unique spectral properties in the cytochrome P450s is a magnet for bioinorganic chemists and biophysicists. The \ndifficult conversion of unactivated hydrocarbons attracted the bioorganic chemist. With the genome revolution and insights into the complex \nprocess of transcriptional and translational regulation, biochemists and molecular biologists found exciting problems in the study of CYPs.\n\nA continuing challenge is to understand how the diverse set of substrate specificities and metabolic transformations are determined by the \nprecise nature of the heme−iron oxygen and protein structure. The structure and electronic configuration of the active oxygen \nintermediates which serve as efficient catalysts remains an area of active research. Complicating this richness in metabolic potential \nis the importance of genetic differences, including single nucleotide polymorphisms, which can alter the physiological responses of the \ncytochrome P450s. Thus, over the past five−plus decades one has seen the evolution from a whole−organ and animal pharmacology \napproach to a quest for the molecular details necessary for precise understanding of structure and function of the P450 systems in \nmaintaining cellular homeostasis. The P450s are now recognized to occupy a great variety of phylogenetically distributed isoform \nactivities, and these variations in metabolic profile and substrate specificity are ultimately dictated by the bioinorganic chemistry \nof heme iron and oxygen as controlled by the protein environment.\n\nWith the elucidation of precise structures for many P450 hemoproteins as well as the application of varied biochemical and biophysical \nmethodologies, this diverse class of oxygenases is beginning to yield its secrets. Much remains to be learned, however, as many of the \nfundamental chemical entities and catalytic details, though perhaps described in textbooks, are in fact still poorly understood. The \nfocus of this review is to place the current knowledge base of cytochrome P450 structure−function in context with the general \naspects of metalloenzyme function. In 2006 Dr. Hayaishi, the founder of this broad field of oxygen metabolism, will celebrate an \nimportant birthday. Hopefully, in reading this review, he will be struck with the outstanding progress that has been realized with \nthis one particular oxygenase and at the same time perhaps provide some important suggestions as to pathways for solving the remaining problems.4\n\nCytochrome P450 has benefited from the attention of inorganic, organic, and physical chemists since its discovery due to its unique \nspectral properties as well as its ability to efficiently catalyze a variety of difficult biotransformations. With the discovery of \nP450 involvement in steroid biosynthesis in the 1970s, joined with its central function in drug metabolism, with its role in a variety \nof other pharmaceutical applications, P450 became one of the most intensively investigated biochemical systems. Multiple monographs, \nprinted conference proceedings, and thematic books have been published as well as special Methods in Enzymology volumes, only a few \nof which can be referenced here.5−12\n\nThe cytochrome P450s became most known for their efficiency in hydroxylation of unactivated alkanes as only a select few oxygenases \npossess the requisite active oxygen state. With equal efficacy, P450s can carry out a wide variety of biotransformations. The list \nin ref 13 includes more than 20 different chemical reactions. Some more unusual reactions catalyzed by P450 were recently reviewed by Guengerich.14\n\nThe mechanism of P450 is a complex cascade of individual steps involving the interaction of protein redox partners and consumption of \nreducing equivalents, most commonly in the form of NAD(P)H. It is somewhat humbling that the earliest versions of the enzymatic cycle \npublished over 30 years ago had much of the important steps characterized by physical and chemical methods.15 Continual refinement has \nled to more detailed versions and the direct observation and structural characterization of new adducts of iron and oxygen. The current \nversion contains eight intermediates, including highly transient caged radical pairs, and has been reviewed from various perspectives.11,12,16−19\n\nWhile the basic concepts central to P450 catalysis were appreciated by early 1970, notable progress in the detailed understanding of these \nmechanisms has been made in the past decade. This has been possible due to the accumulation of exciting data generated through application \nof a wide set of new methodologies, including systematic directed mutagenesis, high−resolution X−ray crystal structure \ndetermination, multiparametric spectroscopic characterization of intermediates, isolation of critical steps using cryogenic or fast \nkinetic techniques, and many excellent quantum chemical and molecular dynamics computational studies. The current view of the oxygen \nactivation mechanisms, catalyzed by metal centers in heme enzymes (as well as in non−heme enzymes, which lie outside the scope \nof this review), ensures one with a much better opportunity to see the common mechanistic picture than was possible earlier.20 \nSuccessful mechanistic studies of other heme enzymes which use different forms of so−called 'active oxygen \nintermediates', such as peroxidases,21,22 heme oxygenases (HO),23−25 catalases,26 nitric oxide synthases \n(NOS),27,28 peroxygenases,29,30 provide a vision of a highly diverse cofactor. Mechanistic insight from each of these various \nsystems has provided important complementary insight into cytochrome P450 mechanism. A fundamental question remaining is how the \nprotein controls efficient performance of such different functions using similar highly reactive heme−oxygen complexes. The \ncomparison of similar reactive intermediates in different enzymes helps to distinguish between the essential features of each of the \nenzymes and so provides additional clues to the revelation of the active role of the protein in heme−enzyme catalysis. The \nrecent progress in isolation and cryogenic stabilization of some of these intermediates makes possible direct spectroscopic and \nstructural studies of this type.\n\nAn exhaustive review of all achievements in oxygen activation chemistry is clearly difficult, even if the field is limited to \nthe processes directly relevant to P450 catalysis. Discussion of the P450 cat

Citations Over TimeTop 1% of 2005 papers

Abstract

Related Papers