Cytochromes P450 (CYPs) are heme-containing enzymes that catalyze various oxidation reactions. They are found in all domains of life and play important roles in metabolism. In humans, CYPs metabolize many drugs and other xenobiotic compounds in the liver and other tissues. The CYP enzymes transfer electrons from NADPH via electron transport systems to oxygenase substrates. Over 50 CYP genes have been identified in humans, with the gene products categorized into families and subfamilies based on amino acid sequence similarity. CYP enzymes are involved in drug metabolism and are subject to drug interactions that can affect their activity levels.

1. Cytochromes P450 (CYPs) belong to the superfamily of proteins containing a heme cofactor and,
therefore, arehemoproteins. CYPs use a variety of small and large molecules as substrates in
enzymatic reactions. They are, in general, the terminal oxidase enzymes in electron transfer chains,
broadly categorized as P450-containing systems. The term P450 is derived from
the spectrophotometric peak at the wavelength of the absorption maximum of the enzyme (450 nm)
when it is in the reduced state and complexed with CO.
CYP enzymes have been identified in all domains of life -
animals, plants, fungi, protists, bacteria, archaea, and even inviruses.[1]
However, the enzymes have
not been found in E. coli.[2][3]
More than 21,000 distinct CYP proteins are known.[4]
Most CYPs require a protein partner to deliver one or more electrons to reduce the iron (and
eventually molecular oxygen). Based on the nature of the electron transfer proteins, CYPs can be
classified into several groups:[5]
 Microsomal P450 systems in which electrons are transferred from NADPH via cytochrome
P450 reductase(variously CPR, POR, or CYPOR). Cytochrome b5 (cyb5) can also contribute
reducing power to this system after being reduced by cytochrome b5 reductase (CYB5R).
 Mitochondrial P450 systems, that employ adrenodoxin reductase and adrenodoxin to transfer
electrons from NADPH to P450.
 Bacterial P450 systems, that employ a ferredoxin reductase and a ferredoxin to transfer
electrons to P450.
 CYB5R/cyb5/P450 systems in which both electrons required by the CYP come
from cytochrome b5.
 FMN/Fd/P450 systems originally found in Rhodococcus sp. in which a FMN-domain-containing
reductase is fused to the CYP.
 P450 only systems, which do not require external reducing power. Notable ones
include CYP5 (thromboxane synthase), CYP8 (prostacyclin synthase), and CYP74A (allene
oxide synthase).
The most common reaction catalyzed by cytochromes P450 is a monooxygenase reaction, e.g.,
insertion of one atom of oxygen into the aliphatic position of an organic substrate (RH) while the
other oxygen atom is reduced to water:
RH + O2 + NADPH + H+
→ ROH + H2O + NADP+
Contents
[hide]
 1 Nomenclature
 2 Mechanism
 3 P450s in humans

2. o 3.1 Drug metabolism
 3.1.1 Drug interaction
 3.1.2 Interaction of other substances
o 3.2 Other specific CYP functions
o 3.3 CYP families in humans
 4 P450s in other species
o 4.1 Animals
o 4.2 Microbial
o 4.3 Fungi
o 4.4 Plants
 5 P450s in biotechnology
 6 InterPro subfamilies
 7 References
 8 External links
Nomenclature[edit]
Genes encoding CYP enzymes, and the enzymes themselves, are designated with the
abbreviation CYP, followed by a number indicating the gene family, a capital letter indicating the
subfamily, and another numeral for the individual gene. The convention is to italicise the name when
referring to the gene. For example,CYP2E1 is the gene that encodes the enzyme CYP2E1 – one of
the enzymes involved in paracetamol (acetaminophen) metabolism. The CYP nomenclature is the
official naming convention, although occasionally (and incorrectly) CYP450 or CYP450 is used.
However, some gene or enzyme names for CYPs may differ from this nomenclature, denoting the
catalytic activity and the name of the compound used as substrate. Examples
include CYP5A1, thromboxane A2 synthase, abbreviated to TBXAS1 (ThromBoXane A2 Synthase 1),
and CYP51A1, lanosterol 14-α-demethylase, sometimes unofficially abbreviated to LDM according
to its substrate (Lanosterol) and activity (DeMethylation).[6]
The current nomenclature guidelines suggest that members of new CYP families share >40% amino
acid identity, while members of subfamilies must share >55% amino acid identity. There are
nomenclature committees that assign and track both base gene names (Cytochrome P450
Homepage) and allele names (CYP Allele Nomenclature Committee).
Mechanism[edit]
Main article: P450-containing systems

3. The P450 catalytic cycle
The active site of cytochrome P450 contains a heme iron center. The iron is tethered to the P450
protein via a thiolate ligand derived from a cysteine residue. This cysteine and several flanking
residues are highly conserved in known CYPs and have the formal PROSITE signature consensus
pattern [FW] - [SGNH] - x - [GD] - {F} - [RKHPT] - {P} - C - [LIVMFAP] - [GAD].[7]
Because of the vast
variety of reactions catalyzed by CYPs, the activities and properties of the many CYPs differ in many
aspects. In general, the P450 catalytic cycle proceeds as follows:
1. The substrate binds to the active site of the enzyme, in proximity to the heme group, on the
side opposite the peptide chain. The bound substrate induces a change in the conformation
of the active site, often displacing a water molecule from the distal axial coordination position
of the heme iron,[8]
and sometimes changing the state of the heme iron from low-spin to
high-spin.[9]
This gives rise to a change in the spectral properties of the enzyme, with an
increase in absorbance at 390 nm and a decrease at 420 nm. This can be measured by
difference spectrometry and is referred to as the "type I" difference spectrum (see inset
graph in figure). Some substrates cause an opposite change in spectral properties, a
"reverse type I" spectrum, by processes that are as yet unclear. Inhibitors and certain
substrates that bind directly to the heme iron give rise to the type II difference spectrum, with
a maximum at 430 nm and a minimum at 390 nm (see inset graph in figure). If no reducing
equivalents are available, this complex may remain stable, allowing the degree of binding to
be determined from absorbance measurements in vitro[10]
2. The change in the electronic state of the active site favors the transfer of an electron from
NAD(P)H via cytochrome P450 reductase or another associatedreductase.[11]
This takes

4. place by way of the electron transfer chain, as described above, reducing the ferric heme
iron to the ferrous state.
3. Molecular oxygen binds covalently to the distal axial coordination position of the heme iron.
The cysteine ligand is a better electron donor than histidine, which is normally found in
heme-containing proteins. As a consequence, the oxygen is activated to a greater extent
than in other heme proteins. However, this sometimes allows the iron-oxygen bond to
dissociate, causing the so-called "uncoupling reaction", which releases a reactive
superoxide radical and interrupts the catalytic cycle.[8]
4. A second electron is transferred via the electron-transport system, from either cytochrome
P450 reductase, ferredoxins, or cytochrome b5, reducing the dioxygen adduct to a
negatively charged peroxo group. This is a short-lived intermediate state.
5. The peroxo group formed in step 4 is rapidly protonated twice by local transfer from water or
from surrounding amino-acid side-chains, releasing one water molecule, and forming a
highly reactive species commonly referred to as P450 Compound 1 ( or Compound I). This
highly reactive intermediate was not "seen in action" until 2010,[12]
although it had been
studied theoretically for many years.[8]
P450 Compound 1 is most likely an iron(IV)oxo
(or ferryl) species with an additional oxidizing equivalent delocalized over the porphyrin and
thiolate ligands. Evidence for the alternative perferryl iron(V)-oxo [8]
is lacking.[12]
6. Depending on the substrate and enzyme involved, P450 enzymes can catalyze any of a wide
variety of reactions. A hypothetical hydroxylation is shown in this illustration. After the
product has been released from the active site, the enzyme returns to its original state, with
a water molecule returning to occupy the distal coordination position of the iron nucleus.
S: An alternative route for mono-oxygenation is via the "peroxide shunt": Interaction with single-
oxygen donors such as peroxides and hypochlorites can lead directly to the formation of the iron-oxo
intermediate, allowing the catalytic cycle to be completed without going through steps 2, 3, 4, and
5.[10]
A hypothetical peroxide "XOOH" is shown in the diagram.
C: If carbon monoxide (CO) binds to reduced P450, the catalytic cycle is interrupted. This reaction
yields the classic CO difference spectrum with a maximum at 450 nm.
P450s in humans[edit]
Human CYPs are primarily membrane-associated proteins[13]
located either in the inner membrane
of mitochondria or in the endoplasmic reticulum of cells. CYPs metabolize thousands
of endogenous and exogenous chemicals. Some CYPs metabolize only one (or a very few)
substrates, such as CYP19 (aromatase), while others may metabolize multiple substrates. Both of
these characteristics account for their central importance in medicine. Cytochrome P450 enzymes
are present in most tissues of the body, and play important roles in hormone synthesis and
breakdown (including estrogen and testosterone synthesis and metabolism),cholesterol synthesis,

5. and vitamin D metabolism. Cytochrome P450 enzymes also function to metabolize potentially toxic
compounds, including drugs and products of endogenous metabolism such as bilirubin, principally in
the liver.
The Human Genome Project has identified 57 human genes coding for the various cytochrome P450
enzymes.[14]
Drug metabolism[edit]
Proportion of antifungal drugs metabolized by different families ofCYPs.[15]
Further information: Drug metabolism
CYPs are the major enzymes involved in drug metabolism, accounting for about 75% of the total
metabolism.[16]
Most drugs undergo deactivation by CYPs, either directly or by
facilitatedexcretion from the body. Also, many substances are bioactivated by CYPs to form their
active compounds.
Drug interaction[edit]
Many drugs may increase or decrease the activity of various CYP isozymes either by inducing the
biosynthesis of an isozyme (enzyme induction) or by directly inhibiting the activity of the CYP
(enzyme inhibition). This is a major source of adverse drug interactions, since changes in CYP
enzyme activity may affect the metabolism and clearance of various drugs. For example, if one drug
inhibits the CYP-mediated metabolism of another drug, the second drug may accumulate within the
body to toxic levels. Hence, these drug interactions may necessitate dosage adjustments or
choosing drugs that do not interact with the CYP system. Such drug interactions are especially
important to take into account when using drugs of vital importance to the patient, drugs with
important side-effects and drugs with small therapeutic windows, but any drug may be subject to an
altered plasma concentration due to altered drug metabolism.
A classical example includes anti-epileptic drugs. Phenytoin, for example,
induces CYP1A2, CYP2C9, CYP2C19, and CYP3A4. Substrates for the latter may be drugs with

6. critical dosage, like amiodarone or carbamazepine, whose blood plasma concentration may either
increase because of enzyme inhibition in the former, or decrease because of enzyme induction in
the latter.[citation needed]
Interaction of other substances[edit]
Naturally occurring compounds may also induce or inhibit CYP activity. For
example, bioactive compounds found in grapefruit juice and some other fruit juices,
including bergamottin, dihydroxybergamottin, and paradicin-A, have been found to inhibit CYP3A4-
mediated metabolism of certain medications, leading to increasedbioavailability and, thus, the strong
possibility of overdosing.[17]
Because of this risk, avoiding grapefruit juice and fresh grapefruits
entirely while on drugs is usually advised.[18]
Other examples:
 Saint-John's wort, a common herbal remedy induces CYP3A4, but also
inhibits CYP1A1, CYP1B1, and CYP2D6.[19][20]
 Tobacco smoking induces CYP1A2 (example CYP1A2 substrates are clozapine, olanzapine,
and fluvoxamine)[21]
 At relatively high concentrations, starfruit juice has also been shown to inhibit CYP2A6 and other
CYPs.[22]
Watercress is also a known inhibitor of the Cytochrome P450 CYP2E1, which may
result in altered drug metabolism for individuals on certain medications (ex., chlorzoxazone).[23]
 Tributyltin has been found to inhibit the function of Cytochrome P450, leading to masculinization
of mollusks.[24]
Other specific CYP functions[edit]

7. Steroidogenesis,showing manyofthe enzyme activities that are performed by cytochrome P450 enzymes. HSD:
Hydroxysteroid dehydrogenase
A subset of cytochrome P450 enzymes play important roles in the synthesis of steroid
hormones (steroidogenesis) by the adrenals, gonads, and peripheral tissue:
 CYP11A1 (also known as P450scc or P450c11a1) in adrenal mitochondria affects “the activity
formerly known as 20,22-desmolase” (steroid 20α-hydroxylase, steroid 22-hydroxylase,
cholesterol side-chain scission).
 CYP11B1 (encoding the protein P450c11β) found in the inner mitochondrial
membrane ofadrenal cortex has steroid 11β-hydroxylase, steroid 18-hydroxylase, and steroid
18-methyloxidase activities.
 CYP11B2 (encoding the protein P450c11AS), found only in the mitochondria of the adrenalzona
glomerulosa, has steroid 11β-hydroxylase, steroid 18-hydroxylase, and steroid 18-
methyloxidase activities.
 CYP17A1, in endoplasmic reticulum of adrenal cortex has steroid 17α-hydroxylase and 17,20-
lyase activities.
 CYP21A1 (P450c21) in adrenal cortexconducts 21-hydroxylase activity.
 CYP19A (P450arom, aromatase) in endoplasmic reticulum of gonads, brain, adipose tissue, and
elsewhere catalyzes aromatization of androgens to estrogens.
CYP families in humans[edit]

8. Humans have 57 genes and more than 59 pseudogenes divided among 18 families of cytochrome
P450 genes and 43 subfamilies.[25]
This is a summary of the genes and of the proteins they encode.
See the homepage of the Cytochrome P450 Nomenclature Committee for detailed information.[14]
Fa
mi
ly
Function
Memb
ers
Names
C
Y
P1
drug and steroid
(especially estrogen)
metabolism, benzo(a)
pyrene toxification
3
subfa
milies,
3
genes
,
1 pse
udoge
ne
CYP1A1, CYP1A2, CYP1B1
C
Y
P2
drug
and steroid metabolis
m
13
subfa
milies,
16
genes
,
16pse
udoge
nes
CYP2A6, CYP2A7, CYP2A13, CYP2B6, CYP2C8, CYP2C9, CYP
2C18, CYP2C19,CYP2D6, CYP2E1, CYP2F1, CYP2J2, CYP2R1,
CYP2S1, CYP2U1, CYP2W1
C
Y
P3
drug
and steroid (including
testosterone)
metabolism
1
subfa
mily, 4
genes
,
2pseu
dogen
CYP3A4, CYP3A5, CYP3A7, CYP3A43

9. es
C
Y
P4
arachidonic acid or
fatty acid metabolism
6
subfa
milies,
12
genes
,
10pse
udoge
nes
CYP4A11, CYP4A22, CYP4B1, CYP4F2, CYP4F3, CYP4F8, CYP
4F11, CYP4F12,CYP4F22, CYP4V2, CYP4X1, CYP4Z1
C
Y
P5
thromboxane A2 synt
hase
1
subfa
mily, 1
gene
CYP5A1
C
Y
P7
bile acid biosynthesis
7-alpha hydroxylase
of steroid nucleus
2
subfa
milies,
2
genes
CYP7A1, CYP7B1
C
Y
P8
varied
2
subfa
milies,
2
genes
CYP8A1 (prostacyclin synthase), CYP8B1 (bile acid biosynthesis)
C
Y
P1
1
steroid biosynthesis
2
subfa
milies,
3
CYP11A1, CYP11B1, CYP11B2

10. genes
C
Y
P1
7
steroid biosynthesis,
17-alpha hydroxylase
1
subfa
mily, 1
gene
CYP17A1
C
Y
P1
9
steroid biosynthesis:
aromatasesynthesize
s estrogen
1
subfa
mily, 1
gene
CYP19A1
C
Y
P2
0
unknown function
1
subfa
mily, 1
gene
CYP20A1
C
Y
P2
1
steroid biosynthesis
2
subfa
milies,
1
gene,
1
pseud
ogene
CYP21A2
C
Y
P2
4
vitamin
D degradation
1
subfa
mily, 1
gene
CYP24A1
C
Y
retinoic
3
subfa
CYP26A1, CYP26B1, CYP26C1

11. P2
6
acid hydroxylase milies,
3
genes
C
Y
P2
7
varied
3
subfa
milies,
3
genes
CYP27A1 (bile acid biosynthesis), CYP27B1 (vitamin D3 1-alpha
hydroxylase, activates vitamin D3), CYP27C1 (unknown function)
C
Y
P3
9
7-alpha hydroxylation
of 24-
hydroxycholesterol
1
subfa
mily, 1
gene
CYP39A1
C
Y
P4
6
cholesterol 24-
hydroxylase
1
subfa
mily, 1
gene
CYP46A1
C
Y
P5
1
cholesterol biosynthe
sis
1
subfa
mily, 1
gene,
3
pseud
ogene
s
CYP51A1 (lanosterol 14-alpha demethylase)
P450s in other species[edit]
Animals[edit]
Many animals have as many or more CYP genes than humans do. For example, mice have genes
for 101 CYPs, and sea urchins have even more (perhaps as many as 120 genes).[26]
Most CYP

12. enzymes are presumed to have monooxygenase activity, as is the case for most mammalian CYPs
that have been investigated (except for, e.g., CYP19 and CYP5). However, gene and genome
sequencing is far outpacing biochemical characterization of enzymatic function, although many
genes with close homology to CYPs with known function have been found.
The classes of CYPs most often investigated in non-human animals are those either involved
in development (e.g., retinoic acid or hormone metabolism) or involved in the metabolism of toxic
compounds (such as heterocyclic amines or polyaromatic hydrocarbons). Often there are differences
in gene regulation or enzyme function of CYPs in related animals that explain observed differences
in susceptibility to toxic compounds (ex. canines inability to metabolize xanthines such as caffeine).
Some drugs undergo metabolism in both species via different enzymes, resulting in different
metabolites, while other drugs are metabolized in one species but excreted unchanged in another
species. For this reason, one species reaction to a substance is not a reliable indication of the
substances effects in humans.
CYPs have been extensively examined in mice, rats, dogs, and less so in zebrafish, in order to
facilitate use of these model organisms in drug discovery andtoxicology. Recently CYPs have also
been discovered in avian species, in particular turkeys, that may turn out to be a great model for
cancer research in humans.[27]
CYP1A5 and CYP3A37 in turkeys were found to be very similar to the
human CYP1A2 and CYP3A4 respectively, in terms of their kinetic properties as well as in the
metabolism of aflatoxin B1.[28]
CYPs have also been heavily studied in insects, often to understand pesticide resistance. For
example, CYP6G1 is linked to insecticide resistance in DDT-resistantDrosophila melanogaster[29]
and
CYP6Z1 in the mosquito malaria vector Anopheles gambiae is capable of directly
metabolizing DDT.[30]
Microbial[edit]
Microbial cytochromes P450 are often soluble enzymes and are involved in diverse metabolic
processes. In bacteria the distribution of P450s is very variable with many bacteria having no
identified P450s (e.g. E.coli). Some bacteria, predominantly actinomycetes, have numerous P450s
(e.g.,[31][32]
). Those so far identified are generally involved in either biotransformation of xenobiotic
compounds (e.g. CYP105A1 from Streptomyces griseolus metabolizes sulfonylurea herbicides to
less toxic derivatives,[33]
) or are part of specialised metabolite biosynthetic pathways (e.g. CYP170B1
catalyses production of the sesquiterpenoid albaflavenone in Streptomyces albus,[34]
). Although no
P450 has yet been shown to be essential in a microbe, the CYP105 family is highly conserved with a
representative in every streptomycete genome sequenced so far ([35]
). Due to the solubility of
bacterial P450 enzymes, they are generally regarded as easier to work with than the predominantly
membrane bound eukaryotic P450s. This, combined with the remarkable chemistry they catalyse,
has led to many studies using the heterologously expressed proteins in vitro. Few studies have

13. investigated what P450s do in vivo, what the natural substrate(s) are and howP450s contribute to
survival of the bacteria in the natural environment.Three examples that have contributed significantly
to structural and mechanistic studies are listed here, but many different families exist.
 Cytochrome P450cam (CYP101) originally from Pseudomonas putida has been used as a
model for many cytochromes P450 and was the first cytochrome P450 three-dimensional protein
structure solved by X-ray crystallography. This enzyme is part of a camphor-hydroxylating
catalytic cycle consisting of two electron transfer steps from putidaredoxin, a 2Fe-2S cluster-
containing protein cofactor.
 Cytochrome P450 eryF (CYP107A1) originally from the actinomycete
bacterium Saccharopolyspora erythraea is responsible for the biosynthesis of
the antibioticerythromycin by C6-hydroxylation of the macrolide 6-deoxyerythronolide B.
 Cytochrome P450 BM3 (CYP102A1) from the soil bacterium Bacillus megaterium catalyzes the
NADPH-dependent hydroxylation of several long-chain fatty acidsat the ω–1 through ω–3
positions. Unlike almost every other known CYP (except CYP505A1, cytochrome P450 foxy), it
constitutes a natural fusion protein between the CYP domain and an electron donating cofactor.
Thus, BM3 is potentially very useful in biotechnological applications.[36][37]
 Cytochrome P450 119 (CYP119) isolated from
the thermophillic archea Sulfolobus acidocaldarius [38]
has been used in a variety of mechanistic
studies.[12]
Because thermophillic enzymes evolved to function at high temperatures, they tend to
function more slowly at room temperature (if at all) and are therefore excellent mechanistic
models.
Fungi[edit]
The commonly used azole class antifungal drugs work by inhibition of the fungal cytochrome P450
14α-demethylase. This interrupts the conversion of lanosterol toergosterol, a component of the
fungal cell membrane. (This is useful only because humans' P450 have a different sensitivity; this is
how this class of antifungalswork.)[39]
Significant research is ongoing into fungal P450s, as a number of fungi are pathogenic to humans
(such as Candida yeast and Aspergillus) and to plants.
Cunninghamella elegans is a candidate for use as a model for mammalian drug metabolism.
Plants[edit]
Plant cytochromes P450 are involved in a wide range of biosynthetic reactions, leading to
various fatty acid conjugates, plant hormones, defensive compounds, or medically important

14. drugs. Terpenoids, which represent the largest class of characterized natural plant compounds, are
often substrates for plant CYPs.
P450s in biotechnology[edit]
The remarkable reactivity and substrate promiscuity of P450s have long attracted the attention of
chemists.[40]
Recent progress towards realizing the potential of using P450s towards difficult
oxidations have included: (i) eliminating the need for natural co-factors by replacing them with
inexpensive peroxide containing molecules,[41]
(ii) exploring the compatibility of p450s with organic
solvents,[42]
and (iii) the use of small, non-chiral auxiliaries to predictably direct P450 oxidation.[citation
needed]
InterPro subfamilies[edit]
InterPro subfamilies:
 Cytochrome P450, B-class IPR002397
 Cytochrome P450, mitochondrial IPR002399
 Cytochrome P450, E-class, group I IPR002401
 Cytochrome P450, E-class, group II IPR002402
 Cytochrome P450, E-class, group IV IPR002403
 Hydroxylate estrogen (CYP1A2 and 1B1)
Clozapine, imipramine, paracetamol, phenacetin Heterocyclic aryl amines Inducible and CYP1A2 5-
10% deficient oxidize uroporphyrinogen to uroporphyrin (CYP1A2) in heme metabolism, but they
may have additional undiscovered endogenous substrates. are inducible by some polycyclic
hydrocarbons, some of which are found in cigarette smoke and charred food. These enzymes are of
interest, because in assays, they can activate compounds to carcinogens. High levels of CYP1A2
have been linked to an increased risk of colon cancer. Since the 1A2 enzyme can be induced by
cigarette smoking, this links smoking with colon cancer.[citation needed]
References[edit]
1. Jump up^ Lamb DC, Lei L, Warrilow AG, Lepesheva GI, Mullins JG, Waterman MR, Kelly SL
(2009). "The first virally encoded cytochrome P450". Journal of Virology 83 (16): 8266-9. PMID
19515774
2. Jump up^ Roland Sigel; Sigel, Astrid; Sigel, Helmut (2007). The Ubiquitous Roles of
Cytochrome P450 Proteins: Metal Ions in Life Sciences. New York: Wiley. ISBN 0-470-01672-8.
3. Jump up^ Danielson PB (December 2002). "The cytochrome P450 superfamily: biochemistry,
evolution and drug metabolism in humans". Curr. Drug Metab. 3 (6): 561–
97. doi:10.2174/1389200023337054. PMID 12369887.
4. Jump up^ Nelson D. "Cytochrome P450 Homepage". University of Tennessee. Retrieved2014-
11-13.

15. Glutathione (GSH) is an important antioxidant in plants, animals, fungi, and some bacteria and
archaea, preventing damage to important cellular components caused by reactive oxygen
species such as free radicals and peroxides.[2]
It is a tripeptide with a gamma peptide
linkage between the carboxyl group of the glutamate side-chain and the amine
group of cysteine (which is attached by normal peptide linkage to a glycine).
Thiol groups are reducing agents, existing at a concentration around 5 mM in animal cells.
Glutathione reducesdisulfide bonds formed within cytoplasmic proteins to cysteines by serving as
an electron donor. In the process, glutathione is converted to its oxidized form, glutathione
disulfide (GSSG), also called L-(–)-glutathione.
Once oxidized, glutathione can be reduced back by glutathione reductase, using NADPH as an
electron donor.[3]
The ratio of reduced glutathione to oxidized glutathione within cells is often used as
a measure of cellular toxicity.[4]
Contents
[hide]
 1 Biosynthesis
 2 Function
o 2.1 Function in animals
o 2.2 Function in plants
 3 Supplementation
o 3.1 Cancer
 4 Methods to determine glutathione
 5 Importance in winemaking
 6 See also
 7 References
 8 Related research
 9 External links
Biosynthesis[edit]
Glutathione is not an essential nutrient, since it can be synthesized in the body from the amino
acids L-cysteine, L-glutamic acid, and glycine. The sulfhydryl group (SH) of cysteine serves as a
proton donor and is responsible for its biological activity. Cysteine is the rate-limiting factor in cellular
glutathione synthesis, since this amino acid is relatively rare in foods.
Cells make glutathione in two adenosine triphosphate-dependent steps:
 First, gamma-glutamylcysteine is synthesized from L-glutamate and cysteine via the
enzyme gamma-glutamylcysteine synthetase (glutamate cysteine ligase, GCL). This reaction is
the rate-limiting step in glutathione synthesis.[5]

16.  Second, glycine is added to the C-terminal of gamma-glutamylcysteine via the
enzyme glutathione synthetase.
Animal glutamate cysteine ligase (GCL) is a heterodimeric enzyme composed of a catalytic (GCLC)
and a modulatory (GCLM) subunit. GCLC constitutes all the enzymatic activity, whereas GCLM
increases the catalytic efficiency of GCLC. Mice lacking GCLC (i.e., lacking all de novo GSH
synthesis) die before birth.[6]
Mice lacking GCLM demonstrate no outward phenotype, but exhibit
marked decrease in GSH and increased sensitivity to toxic insults.[7][8][9]
While all cells in the human body are capable of synthesizing glutathione, liver glutathione synthesis
has been shown to be essential. Mice with genetically induced loss of GCLC (i.e., GSH synthesis)
only in the liver die within a month of birth.[10]
The plant glutamate cysteine ligase (GCL) is a redox-sensitive homodimeric enzyme, conserved in
the plant kingdom.[11]
In an oxidizing environment, intermolecular disulfide bridges are formed and the
enzyme switches to the dimeric active state. The midpoint potential of the critical cysteine pair is -
318 mV. In addition to the redox-dependent control is the plant GCL enzyme feedback inhibited
by GSH.[12]
GCL is exclusively located in plastids, and glutathione synthetase is dual-targeted to
plastids and cytosol, thus are GSH and gamma-glutamylcysteine exported from the plastids.[13]
Both
glutathione biosynthesis enzymes are essential in plants; knock-outs of GCL and GS are lethal to
embryo and seedling.[14]
The biosynthesis pathway for glutathione is found in some bacteria, such
as cyanobacteria and proteobacteria, but is missing in many other bacteria. Most eukaryotes
synthesize glutathione, including humans, but some do not, such as Leguminosae, Entamoeba,
and Giardia. The only archaea that make glutathione are halobacteria.[15][16]
Function[edit]
Glutathione exists in both reduced (GSH) and oxidized (GSSG) states. In the reduced state, the thiol
group of cysteine is able to donate a reducing equivalent (H+
+ e−
) to other unstable molecules, such
as reactive oxygen species. In donating an electron, glutathione itself becomes reactive, but readily
reacts with another reactive glutathione to form glutathione disulfide (GSSG). Such a reaction is
probable due to the relatively high concentration of glutathione in cells (up to 5 mM in the liver).
GSH can be regenerated from GSSG by the enzyme glutathione reductase (GSR):[3]
NADPH
reduces FAD present in GSR to produce a transient FADH-anion. This anion then quickly breaks a
disulfide bond (Cys58 - Cys63) and leads to Cys63's nucleophilically attacking the nearest sulfide
unit in the GSSG molecule (promoted by His467), which creates a mixed disulfide bond (GS-Cys58)
and a GS-anion. His467 of GSR then protonates the GS-anion to form the first GSH. Next, Cys63
nucleophilically attacks the sulfide of Cys58, releasing a GS-anion, which, in turn, picks up a solvent
proton and is released from the enzyme, thereby creating the second GSH. So, for every GSSG and

17. NADPH, two reduced GSH molecules are gained, which can again act as antioxidants scavenging
reactive oxygen species in the cell.
In healthy cells and tissue, more than 90% of the total glutathione pool is in the reduced form (GSH)
and less than 10% exists in the disulfide form (GSSG). An increased GSSG-to-GSH ratio is
considered indicative of oxidative stress.[17]
Glutathione has multiple functions:
 It is the major endogenous antioxidant produced by the cells, participating directly in the
neutralization of free radicals and reactive oxygen compounds, as well as maintaining
exogenous antioxidants such as vitamins C and E in their reduced (active) forms.[18]
 Regulation of the nitric oxide cycle is critical for life, but can be problematic if unregulated.[19]
 It is used in metabolic and biochemical reactions such as DNA synthesis and repair, protein
synthesis, prostaglandin synthesis, amino acid transport, and enzyme activation. Thus, every
system in the body can be affected by the state of the glutathione system, especially the
immune system, the nervous system, the gastrointestinal system, and the lungs.[citation needed]
 It has a vital function in iron metabolism. Yeast cells depleted of or containing toxic levels of
GSH show an intense iron starvation-like response and impairment of the activity of
extramitochondrial ISC enzymes, followed by death.[20]
Function in animals[edit]
GSH is known as a substrate in both conjugation reactions and reduction reactions, catalyzed
by glutathione S-transferase enzymes in cytosol, microsomes, andmitochondria. However, it is also
capable of participating in nonenzymatic conjugation with some chemicals.
In the case of N-acetyl-p-benzoquinone imine (NAPQI), the reactive cytochrome P450-
reactive metabolite formed by paracetamol (acetaminophen), which becomes toxic when GSH is
depleted by an overdose of acetaminophen, glutathione is an essential antidote to overdose.
Glutathione conjugates to NAPQI and helps to detoxify it. In this capacity, it protects cellular protein
thiol groups, which would otherwise become covalently modified; when all GSH has been spent,
NAPQI begins to react with the cellular proteins, killing the cells in the process. The preferred
treatment for an overdose of this painkiller is the administration (usually in atomized form) of N-
acetyl-L-cysteine (often as a preparation called Mucomyst[21]
), which is processed by cells to L-
cysteine and used in the de novo synthesis of GSH.
Glutathione (GSH) participates in leukotriene synthesis and is a cofactor for the enzyme glutathione
peroxidase. It is also important as a hydrophilic molecule that is added to lipophilic toxins and waste
in the liver during biotransformation before they can become part of the bile. Glutathione is also
needed for the detoxification ofmethylglyoxal, a toxin produced as a byproduct of metabolism.

18. This detoxification reaction is carried out by the glyoxalase system. Glyoxalase I (EC 4.4.1.5)
catalyzes the conversion of methylglyoxal and reduced glutathione toS-D-lactoyl-
glutathione. Glyoxalase II (EC 3.1.2.6) catalyzes the hydrolysis of S-D-lactoyl-glutathione to
glutathione and D-lactic acid.
Glutathione has recently been used as an inhibitor of melanin in the cosmetics industry. In countries
such as Japan and the Philippines, this product is sold as a skin-whitening soap. Glutathione
competitively inhibits melanin synthesis in the reaction of tyrosinase and L-DOPA by interrupting L-
DOPA's ability to bind to tyrosinase during melanin synthesis. The inhibition of melanin synthesis
was reversed by increasing the concentration of L-DOPA, but not by increasing tyrosinase. Although
the synthesized melanin was aggregated within one hour, the aggregation was inhibited by the
addition of glutathione. These results indicate glutathione inhibits the synthesis and agglutination of
melanin by interrupting the function of L-DOPA."[22]
Function in plants[edit]
In plants, glutathione is crucial for biotic and abiotic stress management. It is a pivotal component of
the glutathione-ascorbate cycle, a system that reduces poisonous hydrogen peroxide.[23]
It is the
precursor of phytochelatins, glutathione oligomers that chelate heavy metals such
as cadmium.[24]
Glutathione is required for efficient defence against plant pathogens such
as Pseudomonas syringae and Phytophthora brassicae.[25]
APS reductase, an enzyme of the sulfur
assimilationpathway, uses glutathione as electron donor. Other enzymes using glutathione as
substrate are glutaredoxin, these small oxidoreductases are involved in flower development, salicylic
acid, and plant defence signalling.[26]
Supplementation[edit]
Daily consumption of glutathione is effective at increasing glutathione blood levels. [27]
In a previous study of acute oral administration, seven healthy subjects were given a very large dose
(3 grams) of oral glutathione, Witschi and coworkers found "it is not possible to increase circulating
glutathione to a clinically beneficial extent by the oral administration of a single dose of 3 g of
glutathione." Plasma levels of glutathione, cysteine, and glutamate were tested in the plasma after
270 minutes and three of four of the subjects did have an increase or rise in their glutathione
levels.[28]
However, it is possible to increase and maintain appropriate glutathione levels by
increasing the daily consumption of glutathione-rich foods and/or supplements.[non-primary source needed][29]
Calcitriol (1,25-dihydroxyvitamin D3), the active metabolite of vitamin D3, after being synthesized
from calcifediol in the kidney, increases glutathione levels in the brain and appears to be a catalyst
for glutathione production.[30]
Calcitriol was found to increase GSH levels in rat astrocyte primary
cultures on average by 42%, increasing protein concentrations from 29 nmol/mg to 41 nmol/mg, 24

19. and 48 hours after administration; this effect was reduced to 11%, relative to the control, 96 hours
after administration.[31]
It takes about ten days for the body to process vitamin D3 into calcitriol.[32]
Other supplements, including N-acetylcysteine,[33]
S-adenosylmethionine (SAMe)[34][35][36]
and whey
protein[37][38][39][40]
have also been shown to increase cellular glutathione content in persons suffering
from a disease-related glutathione deficiency.
Zinc has been shown to have some influence over de novo glutathione synthesis.[41][42][43]
Further,
magnesium has significant influence on glutathione synthesis; it appears to be an essential
cofactor.[44][45][46][47][48]
Low glutathione is commonly observed in wasting and negative nitrogen balance,[49]
as seen in
cancer, HIV/AIDS, sepsis, trauma, burns, and athletic overtraining.
Cancer[edit]
Once a tumor has been established, elevated levels of glutathione may act to protect cancerous
cells by conferring resistance to chemotherapeutic drugs.[50]
Methods to determine glutathione[edit]
Reduced glutathione may be visualized using Ellman's reagent or bimane derivatives such
as monobromobimane. The monobromobimane method is more sensitive. In this procedure, cells
are lysed and thiols extracted using a HCl buffer. The thiols are then reduced with dithiothreitol and
labelled by monobromobimane. Monobromobimane becomes fluorescent after binding to GSH. The
thiols are then separated by HPLC and the fluorescence quantified with a fluorescence detector.
Bimane may also be used to quantify glutathione in vivo. The quantification is done by confocal laser
scanning microscopy after application of the dye to living cells.[51]
Another approach, which allows to
measure the glutathione redoxpotential at a high spatial and temporal resolution in living cells is
based on redoximaging using the redox-sensitive green fluorescent protein (roGFP)[52]
or redox
sensitive yellow fluorescent protein (rxYFP)[53]
Importance in winemaking[edit]
The content of glutathione in must determines the browning effect during the production of white
wine by trapping the caffeoyltartaric acid quinones generated by enzymic oxidation as grape reaction
product.[54]
See also[edit]
 Glutathione synthetase deficiency
 Ophthalmic acid
 roGFP, a tool to measure the cellular glutathione redoxpotential

20.  Glutathione-ascorbate cycle
 Bacterial glutathione transferase
 Thioredoxin, a cysteine-containing small proteins with very similar functions as reducing agents
 Glutaredoxin, an antioxidant protein that uses reduced glutathione as a cofactor and is reduced
nonenzymatically by it
 Bacillithiol
 Mycothiol
References[edit]
1. ^ Jump up to:a b Merck Index, 11th Edition, 4369
2. Jump up^ Pompella, A; Visvikis, A; Paolicchi, A; Tata, V; Casini, AF (2003). "The changing
faces of glutathione, a cellular protagonist". Biochemical Pharmacology 66 (8): 1499–
503. doi:10.1016/S0006-2952(03)00504-5. PMID 14555227.
3. ^ Jump up to:a b Couto, Narciso; Malys, Naglis; Gaskell, Simon; Barber, Jill (2013). "Partition and
Turnover of Glutathione Reductase from Saccharomyces cerevisiae: a Proteomic
Approach". Journal of Proteome Research 12 (6): 2885–
94.doi:10.1021/pr4001948. PMID 23631642.
4. Jump up^
2. What is Glutathione?
3. Glutathione (GSH) is often referred to as the body's master antioxidant. Composted of three amino
acids - cysteine, glycine, and glutamate - glutathione can be found in virtually every cell of the
human body. The highest concentration of glutathione is in the liver, making it critical in the body's
detoxification process.
4. Glutathione is also an essential component to the body's natural defense system. Viruses, bacteria,
heavy metal toxicity, radiation, certain medications, and even the normal process of aging can all
cause free-radical damage to healthy cells and deplete glutathione. Glutathione depletion has been
correlated with lower immune function and increased vulnerability to infection due to the liver's
reduced ability to detoxify.
5. As the generation of free radicals exceeds the body's ability to neutralize and eliminate them,
oxidative stress occurs. A primary function of glutathione is to alleviate this oxidative stress.
6. Biochemistry, Metabolism
7. Reduced glutathione (GSH) is a linear tripeptide of L-glutamine, L-cysteine, and glycine. Technically
N-L-gamma-glutamyl-cysteinyl glycine or L-glutathione, the molecule has a sulfhydryl (SH) group
on the cysteinyl portion, which accounts for its strong electron-donating character.
8. As electrons are lost, the molecule becomes oxidized, and two such molecules become linked
(dimerized) by a disulfide bridge to form glutathione disulfide or oxidized glutathione (GS SG). This
linkage is reversible upon re-reduction.
9. GSH is under tight homeostatic control both intracellularly and extracellularly. A dynamic balance is
maintained between GSH synthesis, it’s recycling from GSSG/oxidized glutathione, and its
utilization.
10. GSH synthesis involves two closely linked, enzymatically-controlled reactions that utilize ATP. First,
cysteine and glutamate are combined by gamma-glutamyl cysteinyl synthetase. Second, GSH
synthetase combines gamma-glutamylcysteine with glycine to generate GSH. As GSH levels rise,
they self-limit further GSH synthesis; otherwise, cysteine availability is usually rate-limiting.
Fasting, protein-energy malnutrition, or other dietary amino acid deficiencies limit GSH synthesis.
11. GSH recycling is catalyzed by glutathione disulfide reductase, which uses reducing equivalents from
NADPH to reconvert GSSG to 2GSH. The reducing power of ascorbate helps conserve systemic GSH.

21. 12. GSH is used as a cofactor by (1) multiple peroxidase enzymes, to detoxify peroxides generated from
oxygen radical attack on biological molecules; (2) transhydrogenases, to reduce oxidized centers on
DNA, proteins, and other biomolecules; and (3) glutathione S-transferases (GST) to conjugate GSH
with endogenous substances (e.g., estrogens), exogenous electrophiles (e.g., arene oxides,
unsaturated carbonyls, organic halides), and diverse xenobiotics. Low GST activity may increase risk
for disease—but paradoxically, some GSH conjugates can also be toxic.
13. Direct attack by free radicals and other oxidative agents can also deplete GSH. The homeostatic
glutathione redox cycle attempts to keep GSH repleted as it is being consumed. Amounts available
from foods are limited (less that 150 mg/day), and oxidative depletion can outpace synthesis.
14. The liver is the largest GSH reservoir. The parenchymal cells synthesize GSH for P450 conjugation
and numerous other metabolic requirements—then export GSH as a systemic source of SH-reducing
power. GSH is carried in the bile to the intestinal luminal compartment. Epithelial tissues of the
kidney tubules, intestinal lining and lung have substantial P450 activity—and modest capacity to
export GSH.
15. GSH equivalents circulate in the blood predominantly as cystine, the oxidized and more stable form
of cysteine. Cells import cystine from the blood, reconvert it to cysteine (likely using ascorbate as
cofactor), and from it synthesize GSH. Conversely, inside the cell, GSH helps re -reduce oxidized
forms of other antioxidants—such as ascorbate and alpha-tocopherol.
16. Mechanism of Action
17. GSH is an extremely important cell protectant. It directly quenches reactive hydroxyl free radicals,
other oxygen-centered free radicals, and radical centers on DNA and other biomolecules. GSH is a
primary protectant of skin, lens, cornea, and retina against radiation damage and other biochemical
foundations of P450 detoxification in the liver, kidneys, lungs, intestinal, epithelia and other organs.
18. GSH is the essential cofactor for many enzymes that require thiol-reducing equivalents, and helps
keep redox-sensitive active sites on enzyme in the necessary reduced state. Higher-order thiol cell
systems, the metallothioneins, thioredoxins and other redox regulator proteins are ultimately
regulated by GSH levels—and the GSH/GSSG redox ratio. GSH/GSSG balance is crucial to
homeostasis—stabilizing the cellular biomolecular spectrum, and facilitating cellular performance
and survival.
19. GSH and its metabolites also interface with energetics and neurotransmitter syntheses through
several prominent metabolic pathways. GSH availability down-regulates the pro-inflammatory
potential of leukotrienes and other eicosanoids. Recently discovered S -nitroso metabolites,
generated in vivo from GSH and NO (nitric oxide), further diversify GSH's impact on metabolism.