THE CONCISE GUIDE TO PHARMACOLOGY 2017/18: Enzymes

The Concise Guide to PHARMACOLOGY 2017/18 provides concise overviews of the key properties of nearly 1800 human drug targets with an emphasis on selective pharmacology (where available), plus links to an open access knowledgebase of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide represents approximately 400 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point‐in‐time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.13877/full. Enzymes are one of the eight major pharmacological targets into which the Guide is divided, with the others being: G protein‐coupled receptors, ligand‐gated ion channels, voltage‐gated ion channels, other ion channels, nuclear hormone receptors, catalytic receptors and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid‐2017, and supersedes data presented in the 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature Committee of the Union of Basic and Clinical Pharmacology (NC‐IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

mote a particular conformational change. Co-factors are tightly bound to the enzyme and include metal ions and heme groups. Co-enzymes are typically small molecules which accept or donate functional groups to assist in the enzymatic reaction. Examples include ATP, NAD, NADP and S-adenosylmethionine, as well as a number of vitamins, such as riboflavin (vitamin B1) and thiamine (vitamin B2). Where co-factors/co-enzymes have been identified, the Guide indicates their involvement.

Comments -
The JAK2 V617F mutation, which causes constitutive activation, plays an oncogenic role in the pathogenesis of the myeloproliferative disorders, polycythemia vera, essential thrombocythemia, and idiopathic myelofibrosis [64,115]. Small molecule compounds which inhibit aberrant JAK2 activity are being developed as novel anti-cancer pharmaceuticals.
-- Since it is beyond the scope of the Guide to list all peptidase and proteinase activities, this summary focuses on selected enzymes of significant pharmacological interest that have ligands (mostly small-molecules) directed against them. For those interested in detailed background we recommend the MEROPS database [450] (with whom we collaborate) as an information resource [432].

A22: Presenilin
Enzymes → Peptidases and proteinases → AD: Aspartic (A) Peptidases → A22: Presenilin Overview: Presenilin (PS)-1 or -2 act as the catalytic component/essential co-factor of the γ-secretase complex responsible for the final carboxy-terminal cleavage of amyloid precursor protein (APP) [260] in the generation of amyloid beta (Aβ) [7,510]. Given that the accumulation and aggregation of Aβ in the brain is piv-otal in the development of Alzheimer's disease (AD), inhibition of PS activity is one mechanism being investigated as a therapeutic option for AD [187].

C14: Caspase
Enzymes → Peptidases and proteinases → CD: Cysteine (C) Peptidases → C14: Caspase Overview: Caspases, (E.C. 3.4.22.-) which derive their name from Cysteine ASPartate-specific proteASES, include at least two families; initiator caspases (caspases 2, 8, 9 and 10), which are able to hydrolyse and activate a second family of effector cas-pases (caspases 3, 6 and 7), which themselves are able to hydrolyse further cellular proteins to bring about programmed cell death. Caspases are heterotetrameric, being made up of two pairs of subunits, generated by a single gene product, which is prote-olysed to form the mature protein. Members of the mammalian inhibitors of apoptosis proteins (IAP) are able to bind the procaspases, thereby preventing maturation to active proteinases.
Information on members of this family may be found in the online database.

M1: Aminopeptidase N
Enzymes → Peptidases and proteinases → MA: Metallo (M) Peptidases → M1: Aminopeptidase N Overview: Aminopeptidases catalyze the cleavage of amino acids from the amino (N) terminus of protein or peptide substrates, and are involved in many essential cellular functions. Members of this enzyme family may be monomeric or multi-subunit complexes, and many are zinc metalloenzymes [522].
Information on members of this family may be found in the online database.

Nomenclature
Angiotensin
Information on members of this family may be found in the online database.

Comments
Folate hydrolase is also known as NAALADase as it is responsible for the hydrolysis of N-acetaspartylglutamate to form N-acetylaspartate and L-glutamate (L-glutamic acid). In the gut, the enzyme assists in the assimilation of folate by hydrolysing dietary poly-gamma-glutamylfolate. The enzyme is highly expressed in the prostate, and its expression is up-regulated in cancerous tissue. A tagged version of the antibody capromab has been used for imaging purposes.
Comments: Folate hydrolase is also known as NAALADase as it is responsible for the hydrolysis of N-acetaspartylglutamate to form N-acetylaspartate and L-glutamate. In the gut, the enzyme assists in the assimilation of folate by hydrolysing dietary poly-gamma-glutamylfolate. The enzyme is highly expressed in the prostate, and its expression is up-regulated in cancerous tissue. A tagged version of the antibody capromab has been used for imaging purposes.

T1: Proteasome
Enzymes → Peptidases and proteinases → PB: Threonine (T) Peptidases → T1: Proteasome Overview: The T1 macropain beta subunits form the catalytic proteinase core of the 20S proteasome complex [93]. This catalytic core enables the degradation of peptides with Arg, Phe, Tyr, Leu, and Glu adjacent to the cleavage site. The β5 subunit is the principal target of the approved drug proteasome inhibitor bortezomib.

Acetylcholine turnover
Enzymes → Acetylcholine turnover Overview: Acetylcholine is familiar as a neurotransmitter in the central nervous system and in the periphery. In the somatic nervous system, it activates nicotinic acetylcholine receptors at the skeletal neuromuscular junction. It is also employed in the autonomic nervous system, in both parasympathetic and sympathetic branches; in the former, at the smooth muscle neuromus-cular junction, activating muscarinic acetylcholine receptors. In the latter, acetylcholine is involved as a neurotransmitter at the ganglion, activating nicotinic acetylcholine receptors. Acetylcholine is synthesised in neurones through the action of choline O-acetyltransferase and metabolised after release through the extracellular action of acetylcholinesterase and cholinesterase.
Choline is accumulated from the extracellular medium by selective transporters (see SLC5A7 and the SLC44 family). Acetylcholine is accumulated in synaptic vesicles through the action of the vesicular acetylcholine transporter SLC18A3. Comments Splice variants of choline O-acetyltransferase are suggested to be differentially distributed in the periphery and CNS (see [30]).

--
Comments: A number of organophosphorus compounds inhibit acetylcholinesterase and cholinesterase irreversibly, including pesticides such as chlorpyrifos-oxon, and nerve agents such as tabun, soman and sarin. AChE is unusual in its exceptionally high turnover rate which has been calculated at 740 000/min/molecule [570].

Adenosine turnover
Enzymes → Adenosine turnover Overview: A multifunctional, ubiquitous molecule, adenosine acts at cell-surface G protein-coupled receptors, as well as numerous enzymes, including protein kinases and adenylyl cyclase. Extracellular adenosine is thought to be produced either by export or by metabolism, predominantly through ecto-5'-nucleotidase activity (also producing inorganic phosphate). It is inactivated either by extracellular metabolism via adenosine deaminase (also producing ammonia) or, following uptake by nucleoside trans-porters, via adenosine deaminase or adenosine kinase (requiring ATP as co-substrate). Intracellular adenosine may be produced by cytosolic 5'-nucleotidases or through S-adenosylhomocysteine hydrolase (also producing L-homocysteine).

Nomenclature
Adenosine αβ-methyleneADP (pIC 50 8.7) [56] 3-deazaadenosine (pIC 50 8.5) [197] Comments -The enzyme exists in two isoforms derived from alternative splicing of a single gene product: a short isoform, ADK-S, located in the cytoplasm is responsible for the regulation of intra-and extracellular levels of adenosine and hence adenosine receptor activation; a long isoform, ADK-L, located in the nucleus contributes to the regulation of DNA methylation [48,569].

Amino acid hydroxylases
Enzymes → Amino acid hydroxylases Overview: The amino acid hydroxylases (monooxygenases), EC.1.14.16.-, are iron-containing enzymes which utilise molecular oxygen and sapropterin as co-substrate and co-factor, respectively. In humans, as well as in other mammals, there are two distinct L-Tryptophan hydroxylase 2 genes. In humans, these genes are located on chromosomes 11 and 12 and encode two different homologous enzymes, TPH1 and TPH2.

L-Arginine turnover
Enzymes → L-Arginine turnover Overview: L-arginine is a basic amino acid with a guanidino sidechain. As an amino acid, metabolism of L-arginine to form L-ornithine, catalysed by arginase, forms the last step of the urea production cycle. L-Ornithine may be utilised as a precursor of polyamines (see Carboxylases and Decarboxylases) or recycled via L-argininosuccinic acid to L-arginine. L-Arginine may itself be decarboxylated to form agmatine, although the prominence of this pathway in human tissues is uncertain. L-Arginine may be used as a precursor for guanidoacetic acid formation in the creatine synthesis pathway under the influence of arginine:glycine amidinotransferase with L-ornithine as a byproduct. Nitric oxide synthase uses L-arginine to generate nitric oxide, with L-citrulline also as a byproduct. L-Arginine in proteins may be subject to post-translational mod-ification through methylation, catalysed by protein arginine methyltransferases. Subsequent proteolysis can liberate asymmetric N G ,N G -dimethyl-L-arginine (ADMA), which is an endogenous inhibitor of nitric oxide synthase activities. ADMA is hydrolysed by dimethylarginine dimethylhydrolase activities to generate L-citrulline and dimethylamine. They are dimeric or tetrameric enzymes which use S-adenosyl methionine as a methyl donor, generating S-adenosylhomocysteine as a by-product. They generate both mono-methylated and di-methylated products; these may be symmetric (SDMA) or asymmetric (N G ,N G -dimethyl-L-arginine) versions, where both guanidine nitrogens are monomethylated or one of the two is dimethylated, respectively.
Information on members of this family may be found in the online database.

Arginase
Enzymes → L-Arginine turnover → Arginase Overview: Arginase (EC 3.5.3.1) are manganese-containing isoforms, which appear to show differential distribution, where the ARG1 isoform predominates in the liver and erythrocytes, while ARG2 is associated more with the kidney.
Information on members of this family may be found in the online database.

Nitric oxide synthases
Enzymes → L-Arginine turnover → Nitric oxide synthases Overview: Nitric oxide synthases (NOS, E.C. 1.14.13.39) are a family of oxidoreductases that synthesize nitric oxide (NO.) via the NADPH and oxygen-dependent consumption of L-arginine with the resultant by-product, L-citrulline. There are 3 NOS isoforms and they are related by their capacity to produce NO, highly conserved organization of functional domains and significant homology at the amino acid level. NOS isoforms are functionally distinguished by the cell type where they are expressed, intracellular targeting and transcriptional and post-translation mechanisms regulating enzyme activity. The nomenclature suggested by NC-IUPHAR of NOS I, II and III [363] has not gained wide acceptance, and the 3 isoforms are more commonly referred to as neuronal NOS (nNOS), inducible NOS (iNOS) and endothelial NOS (eNOS) which reflect the location of expression (nNOS and eNOS) and inducible expression (iNOS). All are dimeric enzymes that shuttle electrons from NADPH, which binds to a Cterminal reductase domain, through the flavins FAD and FMN to the oxygenase domain of the other monomer to enable the BH4dependent reduction of heme bound oxygen for insertion into the substrate, L-arginine. Electron flow from reductase to oxygenase domain is controlled by calmodulin binding to canonical calmodulin binding motif located between these domains. eNOS and nNOS isoforms are activated at concentrations of calcium greater than 100 nM, while iNOS shows higher affin-ity for Ca 2+ /calmodulin (CALM1 CALM2 CALM3, P62158) with great avidity and is essentially calcium-independent and constitutively active. Efficient stimulus-dependent coupling of nNOS and eNOS is achieved via subcellular targeting through respective N-terminal PDZ and fatty acid acylation domains whereas iNOS is largely cytosolic and function is independent of intracellular location. nNOS is primarily expressed in the brain and neuronal tissue, iNOS in immune cells such as macrophages and eNOS in the endothelial layer of the vasculature although exceptions in other cells have been documented. L-NAME and related modified arginine analogues are inhibitors of all three isoforms, with IC 50 values in the micromolar range. 3-bromo-7NI (pIC 50 6.1-6.5) [43], 7NI (pIC 50

5.3) [20]
Comments: The reductase domain of NOS catalyses the reduction of cytochrome c and other redox-active dyes [345]. NADPH:O 2 oxidoreductase catalyses the formation of superoxide anion/H 2 O 2 in the absence of L-arginine and sapropterin.

Carboxylases and decarboxylases
Enzymes → Carboxylases and decarboxylases Propionyl-CoA carboxylase is able to function in both forward and reverse activity modes, as a ligase (carboxylase) or lyase (decarboxylase), respectively.
Loss-of-function mutations in γ-glutamyl carboxylase are associated with clotting disorders.
Comments: Dicarboxylic acids including citric acid are able to activate ACC1/ACC2 activity allosterically. PCC is able to function in forward and reverse modes as a ligase (carboxylase) or lyase (decarboxylase) activity, respectively. Loss-of-function mutations in GGCX are associated with clotting disorders.

Enzymes → Carboxylases and decarboxylases → Decarboxylases
Overview: The decarboxylases generate CO 2 and the indicated products from acidic substrates, requiring pyridoxal phosphate or pyruvic acid as a co-factor.

Nomenclature
Glutamic Comments L-aspartic acid is a less rapidly metabolised substrate of mouse brain glutamic acid decarboxylase generating βalanine [577]. Autoantibodies against GAD1 and GAD2 are elevated in type 1 diabetes mellitus and neurological disorders (see Further reading). Comments The presence of a functional ADC activity in human tissues has been questioned [96].
AADC is a homodimer. Inhibited by AMP-activated protein kinase-evoked phosphorylation [451] The activity of ODC is regulated by the presence of an antizyme (ENSG00000104904) and an ODC antizyme inhibitor (ENSG00000155096).

Catecholamine turnover
Enzymes → Catecholamine turnover Overview: Catecholamines are defined by the presence of two adjacent hydroxyls on a benzene ring with a sidechain containing an amine.
These hormone/transmitters are synthesized by sequential metabolism from L-phenylalanine via L-tyrosine. Hydroxylation of L-tyrosine generates levodopa, which is decarboxylated to form dopamine. Hydroxylation of the ethylamine sidechain generates (-)-noradrenaline (norepinephrine), which can be methylated to form (-)-adrenaline (epinephrine). In particular neuronal and adrenal chromaffin cells, the catecholamines dopamine, (-)-noradrenaline and (-)-adrenaline are accumulated into vesicles under the influence of the vesicular monoamine transporters (VMAT1/SLC18A1 and VMAT2/SLC18A2). After release into the synapse or the blood-stream, catecholamines are accumulated through the action cell-surface transporters, primarily the dopamine (DAT/SLC6A3) and norepinephrine transporter (NET/SLC6A2). The primary routes of metabolism of these catecholamines are oxidation via monoamine oxidase activities of methylation via catechol O-methyltransferase.

Nomenclature
L-Phenylalanine hydroxylase TH is a homotetramer, which is inhibited by dopamine and other catecholamines in a physiological negative feedback pathway [109].
DBH is a homotetramer. A protein structurally-related to DBH (MOXD1, Q6UVY6) has been described and for which a function has yet to be identified [76].
AADC is a homodimer.

Ceramide turnover
Enzymes → Ceramide turnover Overview: Ceramides are a family of sphingophospholipids synthesized in the endoplasmic reticulum, which mediate cell stress responses, including apoptosis, autophagy and senescence, Serine palmitoyltransferase generates 3-ketosphinganine, which is reduced to sphinganine (dihydrosphingosine). N-Acylation allows the formation of dihydroceramides, which are subsequently reduced to form ceramides. Once synthesized, ceramides are trafficked from the ER to the Golgi bound to the ceramide transfer protein, CERT (COL4A3BP, Q9Y5P4). Ceramide can be metabolized via multiple routes, ensuring tight regulation of its cellular levels. Addition of phosphocholine generates sphingomyelin while carbohydrate is added to form glucosyl-or galactosylce-ramides. Ceramidase re-forms sphingosine or sphinganine from ceramide or dihydroceramide. Phosphorylation of ceramide generates ceramide phosphate. The determination of accurate kinetic parameters for many of the enzymes in the sphingolipid metabolic pathway is complicated by the lipophilic nature of the substrates.

Ceramide synthase
Enzymes → Ceramide turnover → Ceramide synthase Overview: This family of enzymes, also known as sphingosine N-acyltransferase, is located in the ER facing the cytosol with an as-yet undefined topology and stoichiometry. Ceramide synthase in vitro is sensitive to inhibition by the fungal derived toxin, fumonisin B1.

Sphingomyelin synthase
Enzymes → Ceramide turnover → Sphingomyelin synthase Overview: Following translocation from the ER to the Golgi under the influence of the ceramide transfer protein, sphingomyelin synthases allow the formation of sphingomyelin by the transfer of phosphocholine from the phospholipid phosphatidylcholine. Sphingomyelin synthase-related protein 1 is structurally related but lacks sphingomyelin synthase activity. Comments Glycoceramides are an extended family of sphingolipids, differing in the content and organization of the sugar moieties, as well as the acyl sidechains.

Acid ceramidase
Enzymes → Ceramide turnover → Acid ceramidase Overview: The six human ceramidases may be divided on the basis of pH optimae into acid, neutral and alkaline ceramidases, which also differ in their subcellular location.

Neutral ceramidases
Enzymes → Ceramide turnover → Neutral ceramidases Overview: The six human ceramidases may be divided on the basis of pH optimae into acid, neutral and alkaline ceramidases, which also differ in their subcellular location.

Comments
The enzyme is associated with the plasma membrane [516].

-
Comments: ASAH2B appears to be an enzymatically inactive protein, which may result from gene duplication and truncation.

Alkaline ceramidases
Enzymes → Ceramide turnover → Alkaline ceramidases Overview: The six human ceramidases may be divided on the basis of pH optimae into acid, neutral and alkaline ceramidases, which also differ in their subcellular location. Comments: A ceramide kinase-like protein has been identified in the human genome (CERKL, Q49MI3).

Further reading on Ceramide turnover
Aburasayn H et al.

Chromatin modifying enzymes
Enzymes → Chromatin modifying enzymes

Overview:
Chromatin modifying enzymes, and other chromatin-modifying proteins, fall into three broad categories: writers, readers and erasers. The function of these proteins is to dynamically maintain cell identity and regulate processes such as differentiation, development, proliferation and genome integrity via recognition of specific 'marks' (covalent post-translational modifications) on histone proteins and DNA [280]. In normal cells, tissues and organs, precise co-ordination of these proteins ensures expression of only those genes required to specify phenotype or which are required at specific times, for specific functions. Chromatin modifications allow DNA modifications not coded by the DNA sequence to be passed on through the genome and underlies heritable phenomena such as X chromosome inactivation, aging, heterochromatin formation, reprogramming, and gene silencing (epigenetic control). To date at least eight distinct types of modifications are found on histones. These include small covalent modifications such as acetylation, methylation, and phosphorylation, the attachment of larger modifiers such as ubiquitination or sumoylation, and ADP ribosylation, proline isomerization and deimination. Chromatin modifications and the functions they regulate in cells are reviewed by Kouzarides (2007) [280]. Writer proteins include the histone methyltransferases, histone acetyltransferases, some kinases and ubiquitin ligases. Readers include proteins which contain methyl-lysinerecognition motifs such as bromodomains, chromodomains, tudor domains, PHD zinc fingers, PWWP domains and MBT domains. Erasers include the histone demethylases and histone deacetylases (HDACs and sirtuins). Dysregulated epigenetic control can be associated with human diseases such as cancer [137], where a wide variety of cellular and protein abberations are known to perturb chromatin structure, gene transcription and ultimately cellular pathways [27,477]. Due to the reversible nature of epigenetic modifications, chromatin regulators are very tractable targets for drug discovery and the development of novel therapeutics. Indeed, small molecule inhibitors of writers (e.g. azacitidine and decitabine target the DNA methyltransferases DNMT1 and DNMT3 for the treatment of myelodysplastic syndromes [175,565]) and erasers (e.g. the HDAC inhibitors vorinostat, romidepsin and belinostat for the treatment of T-cell lymphomas [153,265]) are already being used in the clinic. The search for the next generation of compounds with improved specificity against chromatin-associated proteins is an area of intense basic and clinical research [61]. Current progress in this field is reviewed by Simó-Riudalbas and Esteller (2015) [478]. Class III contains the sirtuins (SIRT1-7) Class IV contains only HDAC11. Classes I, II and IV use Zn + as a co-factor, whereas catalysis by Class III enzymes requires NAD + as a co-factor, and members of this subfamily have ADP-ribosylase activity in addition to protein deacetylase function [456]. HDACs have more general protein deacetylase activity, being able to deacetylate lysine residues in non-histone proteins [90] such as microtubules [233], the hsp90 chaperone [281] and the tumour suppressor p53 [322].

2.1.1.-Protein arginine N-methyltransferases
Dysregulated HDAC activity has been identified in cancer cells and tumour tissues [305,444], making HDACs attractive molecular targets in the search for novel mechanisms to treat cancer [567]. Several small molecule HDAC inhibitors are already approved for clinical use: romidepsin, belinostat, vorinostat, panobinostat, belinostat, valproic acid and tucidinostat. HDACs and HDAC inhibitors currently in development as potential anticancer therapeutics are reviewed by Simó-Riudalbas and Esteller (2015) [478].
Information on members of this family may be found in the online database.

Cyclic nucleotide turnover/signalling
Enzymes → Cyclic nucleotide turnover/signalling Overview: Cyclic nucleotides are second messengers generated by cyclase enzymes from precursor triphosphates and hydrolysed by phosphodiesterases. The cellular actions of these cyclic nucleotides are mediated through activation of protein kinases (cAMP-and cGMP-dependent protein kinases), ion channels (cyclic nucleotide-gated, CNG, and hyperpolarization and cyclic nucleotide-gated, HCN) and guanine nucleotide exchange factors (GEFs, Epac).

Comments:
Nitric oxide has been proposed to inhibit AC5 and AC6 selectively [223], although it is unclear whether this phenomenon is of physiological significance. A soluble adenylyl cyclase has been described (ADCY10, Q96PN6 [54]), unaffected by either Gα or Gβγ subunits, which has been suggested to be a cytoplasmic bicarbonate (pH-insensitive) sensor [82]. It can be inhibited selectively by KH7 (pIC 50 5.0-5.5) [221].

Overview:
The principle endocannabinoids are 2acylglycerol esters, such as 2-arachidonoylglycerol (2AG), and N-acylethanolamines, such as anandamide (Narachidonoylethanolamine, AEA). The glycerol esters and ethanolamides are synthesised and hydrolysed by parallel, independent pathways. Mechanisms for release and re-uptake of endocannabinoids (and related entities) are unclear, although candidates for intracellular transport have been suggested. For the generation of 2-arachidonoylglycerol, the key enzyme involved is diacylglycerol lipase (DGL), whilst several routes for anandamide synthesis have been described, the best characterized of which involves N-acylphosphatidylethanolaminephospholipase D (NAPE-PLD, [476]). A transacylation enzyme which forms N-acylphosphatidylethanolamines has recently been identified as a cytosolic enzyme, PLA2G4E (Q3MJ16) [383].
In vitro experiments indicate that the endocannabinoids are also substrates for oxidative metabolism via cyclooxygenase, lipoxygenase and cytochrome P450 enzyme activities [11,154,488].

N-Acylethanolamine turnover
Enzymes → Endocannabinoid turnover → N-Acylethanolamine turnover Comments NAPE-PLD activity appears to be enhanced by polyamines in the physiological range [311], but fails to transphosphatidylate with alcohols [408] unlike phosphatidylcholine-specific phospholipase D.
-The FAAH2 gene is found in many primate genomes, marsupials, and other distantly related vertebrates, but not a variety of lower placental mammals, including mouse and rat [563].
-Comments: Routes for N-acylethanolamine biosynthesis other than through NAPE-PLD activity have been identified [536].

Eicosanoid turnover
Enzymes → Eicosanoid turnover Overview: Eicosanoids are 20-carbon fatty acids, where the usual focus is the polyunsaturated analogue arachidonic acid and its metabolites. Arachidonic acid is thought primarily to derive from phospholipase A2 action on membrane phosphatidylcholine, and may be re-cycled to form phospholipid through conjugation with coenzyme A and subsequently glycerol derivatives. Oxidative metabolism of arachidonic acid is conducted through three major enzymatic routes: cyclooxygenases; lipoxygenases and cytochrome P450-like epoxygenases, particularly CYP2J2. Isoprostanes are structural analogues of the prostanoids (hence the nomenclature D-, E-, F-isoprostanes and isothromboxanes), which are produced in the presence of elevated free radicals in a non-enzymatic manner, leading to suggestions for their use as biomarkers of oxidative stress. Molecular targets for their action have yet to be defined.

Leukotriene and lipoxin metabolism
Enzymes → Eicosanoid turnover → Leukotriene and lipoxin metabolism Overview: Leukotriene A 4 (LTA 4 ), produced by 5-LOX activity, and lipoxins may be subject to further oxidative metabolism; ω-hydroxylation is mediated by CYP4F2 and CYP4F3, while βoxidation in mitochondria and peroxisomes proceeds in a manner dependent on coenzyme A conjugation. Conjugation of LTA 4 at the 6 position with reduced glutathione to generate LTC 4 occurs under the influence of leukotriene C 4 synthase, with the subsequent formation of LTD 4  Comments: LTA4H is a member of a family of arginyl aminopeptidases (ENSFM00250000001675), which also includes aminopeptidase B (RNPEP, 9H4A4) and aminopeptidase B-like 1 (RNPEPL1, Q9HAU8). Dipeptidase 1 and 2 are members of a family of membrane dipeptidases, which also includes (DPEP3, Q9H4B8) for which LTD 4 appears not to be a substrate.

Further reading on Eicosanoid turnover
Ackermann

GABA turnover
Enzymes → GABA turnover Overview: The inhibitory neurotransmitter γ-aminobutyrate (GABA, 4-aminobutyrate) is generated in neurones by glutamic acid decarboxylase. GAD1 and GAD2 are differentially expressed during development, where GAD2 is thought to subserve a trophic role in early life and is distributed throughout the cytoplasm. GAD1 is expressed in later life and is more associated with nerve terminals [136] where GABA is principally accumulated in vesicles through the action of the vesicular inhibitory amino acid transporter SLC32A1. The role of γ-aminobutyraldehyde dehydrogenase (ALDH9A1) in neurotransmitter GABA synthesis is less clear. Following release from neurons, GABA may interact with either GABA A or GABA B receptors and may be accumu-lated in neurones and glia through the action of members of the SLC6 family of transporters. Successive metabolism through GABA transaminase and succinate semialdehyde dehydrogenase generates succinic acid, which may be further metabolized in the mitochondria in the tricarboxylic acid cycle.

Phosphoinositide-specific phospholipase C
Enzymes → Glycerophospholipid turnover → Phosphoinositide-specific phospholipase C Overview: Phosphoinositide-specific phospholipase C (PLC, EC 3.1.4.11), catalyses the hydrolysis of PIP 2 to IP 3 and 1,2diacylglycerol, each of which have major second messenger functions. Two domains, X and Y, essential for catalytic activity, are conserved in the different forms of PLC. Isoforms of PLC-β are activated primarily by G protein-coupled receptors through members of the G q/11 family of G proteins. The receptor-mediated activation of PLC-γ involves their phosphorylation by receptor tyrosine kinases (RTK) in response to activation of a variety of growth factor receptors and immune system receptors. PLC-1 may represent a point of convergence of signalling via both G protein-coupled and catalytic receptors. Ca 2+ ions are required for catalytic activity of PLC isoforms and have been suggested to be the major physiological form of regulation of PLC-δ activity. PLC has been suggested to be activated non-selectively by the small molecule m3M3FBS [23], although this mechanism of action has been questioned [284]. The aminosteroid U73122 has been described as an inhibitor of phosphoinositide-specific PLC [485], although its selectivity among the isoforms is untested and it has been reported to occupy the H1 histamine receptor [235].
A binding protein for secretory phospholipase A 2 has been identified which shows modest selectivity for sPLA 2 -1B over sPLA 2 -2A, and also binds snake toxin phospholipase A 2 [13]. The binding protein appears to have clearance function for circulating secretory phospholipase A 2 , as well as signalling functions, and is a candidate antigen for idiopathic membraneous nephropathy [29].

Lipid phosphate phosphatases
Enzymes → Glycerophospholipid turnover → Lipid phosphate phosphatases Overview: Lipid phosphate phosphatases, divided into phosphatidic acid phosphatases or lipins catalyse the dephosphorylation of phosphatidic acid (and other phosphorylated lipid derivatives) to generate inorganic phosphate and diacylglycerol. PTEN, a phosphatase and tensin homolog (BZS, MHAM, MMAC1, PTEN1, TEP1) is a phosphatidylinositol 3,4,5-trisphosphate 3-phosphatase which acts as a tumour suppressor by reducing cellular levels of PI 3,4,5-P, thereby toning down activity of PDK1 and PKB. Loss-of-function mutations are frequently identified as somatic mutations in cancers.

Overview:
Phosphatidylinositol may be phosphorylated at either 3-or 4positions on the inositol ring by PI 3-kinases or PI 4-kinases, respectively.

Further reading on Haem oxygenase
Abraham NG et al.

Hydrogen sulphide synthesis
Enzymes → Hydrogen sulphide synthesis Overview: Hydrogen sulfide is a gasotransmitter, with similarities to nitric oxide and carbon monoxide. Although the enzymes indicated below have multiple enzymatic activities, the focus here is the generation of hydrogen sulphide (H 2 S) and the enzymatic characteristics are described accordingly. Cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE) are pyridoxal phosphate (PLP)-dependent enzymes. 3-mercaptopyruvate sulfurtransferase (3-MPST) functions to generate H 2 S; only CAT is PLP-dependent, while 3-MPST is not. Thus, this third pathway is sometimes referred to as PLP-independent. CBS and CSE are predominantly cytosolic enzymes, while 3-MPST is found both in the cytosol and the mitochondria.

Hydrolases
Enzymes → Hydrolases Overview: Listed in this section are hydrolases not accumulated in other parts of the Concise Guide, such as monoacylglycerol lipase and acetylcholinesterase. Pancreatic lipase is the predominant mechanism of fat digestion in the alimentary system; its inhibition is associated with decreased fat absorption. CES1 is present at lower levels in the gut than CES2 (P23141), but predominates in the liver, where it is responsible for the hydrolysis of many aliphatic, aromatic and steroid esters. Hormone-sensitive lipase is also a relatively non-selective esterase associated with steroid ester hydrolysis and triglyceride metabolism, particularly in adipose tissue. Endothelial lipase is secreted from endothelial cells and regulates circulating cholesterol in high density lipoproteins. Information on members of this family may be found in the online database.

Comments:
In vitro analysis suggested IP 3 and IP 4 were poor substrates for SKIP, synaptojanin 1 and synaptojanin 2, but suggested that PIP 2 and PIP 3 were more efficiently hydrolysed [458].

Lanosterol biosynthesis pathway
Enzymes → Lanosterol biosynthesis pathway Overview: Lanosterol is a precursor for cholesterol, which is synthesized primarily in the liver in a pathway often described as the mevalonate or HMG-CoA reductase pathway. The first two steps (formation of acetoacetyl CoA and the mitochondrial generation of (S)-3-hydroxy-3-methylglutaryl-CoA) are also associated with oxidation of fatty acids. --

Comments:
Thymidylate synthetase allows the interconversion of dUMP and dTMP, thereby acting as a crucial step in DNA synthesis. Purine nucleoside phosphorylase allows separation of a nucleoside into the nucleobase and ribose phosphate for nucleotide salvage. Xanthine dehydrogenase generates urate in the purine degradation pathway. Post-translational modifications of xanthine dehydrogenase convert the enzymatic reaction to a xanthine oxidase, allowing the interconversion of hypoxanthine and xanthine, with the production (or consumption) of reactive oxygen species. Ribonucleotide reductases allow the production of deoxyribonucleotides from ribonucleotides.

Thyroid hormone turnover
Enzymes → Thyroid hormone turnover

Overview:
The thyroid hormones triiodothyronine and thyroxine, usually abbreviated as triiodothyronine and T 4 , respectively, are synthesized in the thyroid gland by sequential metabolism of tyrosine residues in the glycosylated homodimeric protein thyroglobulin (TG, P01266) under the influence of the haem-containing protein iodide peroxidase. Iodide peroxidase/TPO is a haem-containing enzyme, from the same structural family as eosinophil peroxidase (EPX, P11678), lactoperoxidase (LPO, P22079) and myeloperoxidase (MPO, P05164). Circulating thyroid hormone is bound to thyroxine-binding globulin (SERPINA7, P05543).

2-oxoglutarate oxygenases
Enzymes → 1.14.11.29 2-oxoglutarate oxygenases Overview: Hypoxia inducible factor (HIF) is a transcriptional complex that is involved in oxygen homeostasis [466]. At normal oxygen levels, the alpha subunit of HIF (HIF-1α) is targeted for degradation by prolyl hydroxylation by the prolyl hydrolxyases PHD proteins 1-3 (HIF-PHs) whch are 2-oxoglutarate oxygenases responsible for the post-translational modification of a specific proline in each of the oxygen-dependent degradation domains of HIF-1α. Hydroxylated HIFs are then targeted for proteasomal degradation via the von Hippel-Lindau ubiquitination complex [245]. Under hypoxic conditions, the hydroxylation reaction is blunted which results in decreased HIF degradation. The surviving HIFs are then available to translocate to the nucleus where they heterodimerize with HIF-1β, effecting increased expression of hypoxia-inducible genes.
HIF-PH enzymes are being investigated as pharmacological targets as their inhibition mimics the hypoxic state and switches on transcription of genes associated with processes such as erythropoiesis and vasculogenesis [151]. Small molecule HIF-PH inhibitors are in clinical trial as novel therapies for the amelioration of anemia associated with chronic kidney disease [50].

poly(ADP-ribose)polymerases
Enzymes → 2.4.2.30 poly(ADP-ribose)polymerases Overview: The Poly ADP-ribose polymerase family is a series of enzymes, where the best characterised members are nuclear proteins which are thought to function by binding to single strand breaks in DNA, allowing the recruitment of repair enzymes by the synthesis of NAD-derived ADP-ribose polymers, which are subsequently degraded by a glycohydrolase (PARG, Q86W56).  Overview: Farnesyltransferase is a member of the prenyltransferases family which also includes geranylgeranyltransferase types I (EC 2.5.1.59) and II (EC 2.5.1.60) [72]. Protein farnesyltransferase catalyses the post-translational formation of a thioether linkage between the C-1 of an isoprenyl group and a cysteine residue fourth from the C-terminus of a protein (ie to the CaaX motif, where 'a' is an aliphatic amino acid and 'X' is usually serine, methionine, alanine or glutamine; leucine for EC 2.5.1.59) [165]. Farnesyltransferase is a dimer, composed of an alpha and beta subunit and requires Mg 2+ and Zn 2+ ions as cofactors. The active site is located between the subunits. Prenylation creates a hydrophobic domain on protein tails which acts as a membrane anchor.
Substrates of the prenyltransferases include Ras, Rho, Rab, other Ras-related small GTP-binding proteins, G-protein γ-subunits, nuclear lamins, centromeric proteins and many proteins involved in visual signal transduction. In relation to the causative association between oncogenic Ras proteins and cancer, farnesyltransferase has become an important mechanistic drug discovery target.
Information on members of this family may be found in the online database. Overview: Histone deacetylases act as erasers of epigenetic acetylation marks on lysine residues in histones. Removal of the acetyl groups facilitates tighter packing of chromatin (heterochromatin formation) leading to transcriptional repression. The histone deacetylase family has been classified in to five subfamilies based on phylogenetic comparison with yeast homologues: Class I contains HDACs 1, 2, 3 and 8 Class IIa contains HDACs 4, 5, 7 and 9 Class IIb contains HDACs 6 and 10 Class III contains the sirtuins (SIRT1-7) Class IV contains only HDAC11. Classes I, II and IV use Zn + as a co-factor, whereas catalysis by Class III enzymes requires NAD + as a co-factor, and members of this subfamily have ADP-ribosylase activity in addition to protein deacetylase function [456]. HDACs have more general protein deacetylase activity, being able to deacetylate lysine residues in non-histone proteins [91] such as microtubules [233], the hsp90 chaperone [281] and the tumour suppressor p53 [322].
Dysregulated HDAC activity has been identified in cancer cells and tumour tissues [305,444], making HDACs attractive molecular targets in the search for novel mechanisms to treat cancer [567]. Several small molecule HDAC inhibitors are already approved for clinical use: romidepsin, belinostat, vorinostat, panobinostat, belinostat, valproic acid and tucidinostat. HDACs and HDAC inhibitors currently in development as potential anticancer therapeutics are reviewed by Simó-Riudalbas and Esteller (2015) [478].
Information on members of this family may be found in the online database.
Information on members of this family may be found in the online database.

RAS subfamily
Enzymes → 3.6.5.2 Small monomeric GTPases → RAS subfamily Overview: The RAS proteins (HRAS, NRAS and KRAS) are small membrane-localised G protein-like molecules of 21 kd. They act as an on/off switch linking receptor and non-receptor tyrosine kinase activation to downstream cytoplasmic or nuclear events. Binding of GTP activates the switch, and hydrolysis of the GTP to GDP inactivates the switch. The RAS proto-oncogenes are the most frequently mutated class of proteins in human cancers. Common mutations compromise the GTP-hydrolysing ability of the proteins causing constitutive activation [495], which leads to increased cell proliferation and decreased apoptosis [598]. Because of their importance in oncogenic transformation these proteins have become the targets of intense drug discovery effort [25].
Information on members of this family may be found in the online database.  Overview: DNA topoisomerases regulate the supercoiling of nuclear DNA to influence the capacity for replication or transcription. The enzymatic function of this series of enzymes involves cutting the DNA to allow unwinding, followed by re-attachment to reseal the backbone. Members of the family are targetted in anti-cancer chemotherapy.