Isocitrate dehydrogenase: Wikis


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Isocitrate dehydrogenase
Isocitrate dehydrogenase of Escherichia coli Complex.jpg
Crystallographic structure of Escherichia coli isocitrate dehydrogenase.[1]
EC number
CAS number 9028-48-2
IntEnz IntEnz view
ExPASy NiceZyme view
MetaCyc metabolic pathway
PRIAM profile
PDB structures
Gene Ontology AmiGO / EGO
isocitrate dehydrogenase (NAD+)
EC number
CAS number 9001-58-5
IntEnz IntEnz view
ExPASy NiceZyme view
MetaCyc metabolic pathway
PRIAM profile
PDB structures
Gene Ontology AmiGO / EGO

Isocitrate dehydrogenase (EC and (EC, also known as IDH, is an enzyme that participates in the citric acid cycle. It catalyzes the third step of the cycle: the oxidative decarboxylation of isocitrate, producing alpha-ketoglutarate (α-ketoglutarate) and CO2 while converting NAD+ to NADH. This is a two-step process, which involves oxidation of isocitrate (a secondary alcohol) to oxalosuccinate (a ketone), followed by the decarboxylation of the carboxyl group beta to the ketone, forming alpha-ketoglutarate. Another isoform of the enzyme catalyzes the same reaction, however this reaction is unrelated to the citric acid cycle, is carried out in the cytosol as well as the mitochondrion and peroxisome[2] and uses NADP+ as a cofactor instead of NAD+.



The following is a list of human isocitrate dehydrogenase isozymes:


NADP+ dependent

Each NADP+-dependent isozyme functions as a homodimer:

isocitrate dehydrogenase 1 (NADP+), soluble
Symbol IDH1
Entrez 3417
HUGO 5382
OMIM 147700
RefSeq NM_005896
UniProt O75874
Other data
EC number
Locus Chr. 2 q32-qter
isocitrate dehydrogenase 2 (NADP+), mitochondrial
Symbol IDH2
Entrez 3418
HUGO 5383
OMIM 147650
RefSeq NM_002168
UniProt P48735
Other data
EC number
Locus Chr. 15 q21-qter

NAD+ dependent

The isocitrate dehydrogenase 3 isozyme is a heterotetramer that is composed of two alpha subunits, one beta subunit, and one gamma subunit:

isocitrate dehydrogenase 3 (NAD+) alpha
Symbol IDH3A
Entrez 3419
HUGO 5384
OMIM 601149
RefSeq NM_005530
UniProt P50213
Other data
EC number
Locus Chr. 15 q25.1-25.2
isocitrate dehydrogenase 3 (NAD+) beta
Symbol IDH3B
Entrez 3420
HUGO 5385
OMIM 604526
RefSeq NM_174855
UniProt O43837
Other data
EC number
Locus Chr. 20 p13
isocitrate dehydrogenase 3 (NAD+) gamma
Symbol IDH3G
Entrez 3421
HUGO 5386
OMIM 300089
RefSeq NM_174869
UniProt P51553
Other data
EC number
Locus Chr. X q28


The overall free energy for this reaction with either isoform is -8.4 kJ/mol. IDH lowers the Km (Michaelis constant) of isocitrate without lowering the Vmax (maximum reaction rate - see Michaelis constant for more details).


The NAD-IDH is composed of 3 subunits, is allosterically regulated, and requires an integrated Mg2+ or Mn2+ ion. The closest homologue that has a known structure is the E. coli NADP-dependent IDH, which has only 2 subunits and a 13% identity and 29% similarity based on the amino acid sequences, making it dissimilar to human IDH and not suitable for close comparison. All the known NADP-IDHs are homodimers.


The IDH step of the citric acid cycle, due to its large negative free energy change, is one of the irreversible reactions in the citric acid cycle, and, therefore, must be carefully regulated to avoid unnecessary depletion of isocitrate (and therefore an accumulation of alpha-ketoglutarate). The reaction is stimulated by the simple mechanisms of substrate availability (isocitrate, NAD+ or NADP+, Mg2+ / Mn2+ ), product inhibition (by NADH (or NADPH outside the citric acid cycle) and alpha-ketoglutarate), and competitive feedback inhibition (by ATP). (ref?)

Catalytic mechanism

Isocitrate + NADP+ Mg2+(metal ion) --> alpha-ketoglutarate + NADPH + H+ + CO2[3][4][5]

Catalytic mechanism of the breakdown of isocitrate into oxalosuccinate, then into a final product of alpha-ketuglutarate. The oxalosuccinate intermediate is hypothetical; it has never been observed in the decarboxylating version of the enzyme.[6]

Steps broken down

Within the citric acid cycle, isocitrate, produced from the isomerization of citrate, undergoes both oxidation and decarboxylation. Using the enzyme Isocitrate Dehydrogenase (IDH), isocitrate is held within its active site by surrounding arginine, tyrosine, asparagine, serine, threonine, and aspartic acid amino acids. The first box shows the overall isocitrate dehydrogenase reaction. The reactants necessary for this enzyme mechanism to work are isocitrate, NAD+/NADP+, and Mn2+ or Mg2+. The products of the reaction are alpha-ketoglutarate, carbon dioxide, and NADH + H+/NADPH + H+.[4] Water molecules are used to help deprotonate the oxygens (O3) of isocitrate.

The second box is Step 1, which is the oxidation of the alpha-C (C#2).[3][4] Oxidation is the first step that isocitrate goes through. In this process,[3] the alcohol group off the alpha-carbon (C#2) is deprotonated and the electrons flow to the alpha-C forming a ketone group and removing a hydride off C#2 using NAD+/NADP+ as an electron accepting cofactor. The oxidation of the alpha-C allows for a position where electrons (in the next step) will be coming down from the carboxyl group and pushing the electrons (making the double bonded oxygen) back up on the oxygen or grabbing a nearby proton off a nearby Lysine amino acid.

The third box is Step 2, which is the decarboxylation of oxalosuccinate. In this step[3][4], the carboxyl group oxygen is deprotonated by a nearby Tyrosine amino acid and those electrons flow down to carbon 2. Carbon dioxide leaves the beta carbon of isocitrate as a leaving group with the electrons flowing to the ketone oxygen off the alpha-C placing a negative charge on the oxygen of the alpha-C and forming an alpha-beta unsaturated double bond between carbons 2 and 3. The lone pair on the alpha-C oxygen picks up a proton from a nearby Lysine amino acid.

The fourth box is Step 3, which is the saturation of the alpha-beta unsaturated double bond between carbons 2 and 3. In this step of the reaction,[3][4] Lysine deprotonates the oxygen off the alpha carbon and the lone pair of electrons on the oxygen of the alpha carbon comes down reforming the ketone double bond and pushing the lone pair (forming the double bond between the alpha and beta carbon) off, picking up a proton from the nearby Tyrosine amino acid.[7] This reaction results in the formation of alpha-ketoglutarate, NADH + H+/NADPH + H+, and CO2.

Mechanism of catalysis

Oxidoreductase step where NAD+ is used to accept a hydride.ref name="nfr154197/32"/>

Looking at the 3-D structures to the left, two Aspartate amino acids are interacting with two adjacent water molecules (w6 and w8) in the Mn2+ isocitrate porcine IDH complex to deprotonate the alcohol off the alpha-Carbon. The oxidation of the alpha-C also takes place in this picture where NAD+ accepts a hydride resulting in oxalosuccinate. Along with the sp3 to sp2 stereochemical change around the alpha-C, there is a ketone group that is formed form the alcohol group. The formation of this ketone double bond allows for resonance to take place as electrons coming down from the leaving carboxylate group move towards the ketone.

Decarboxylation of oxalosuccinate.[7]

The decarboxylation of oxalosuccinate is a key step in the formation of alpha-ketoglutarate. In this reaction, the lone pair on the adjacent Tyrosine oxygen pulls off the proton of the carboxyl group.[7] This carboxyl group is also referred to as the beta subunit of isocitrate. The deprotonation of the carboxyl proton causes the lone pair of electrons to move down making carbon dioxide and separating from oxalosuccinate. The electrons continue to move towards the alpha carbon pushing the double bond electrons (making the ketone) up to pull a proton off an adjacent Lysine residue. An alpha-beta unsaturated double bond results between carbon 2 and three. As you can see in the picture, the green ion represents either Mg2+ or Mn2+, which is a cofactor necessary for this reaction to occur. The metal-ion forms a little complex through ionic interactions with the oxygen atoms on the fourth and fifth carbons (also known as the gamma subunit of isocitrate).

Saturation of the alpha-beta unsaturated double bond.[7]

After carbon dioxide leaves oxalosuccinate in the decarboxylation step, the alcohol on carbon 2 will want to reform the ketone double bond because it is a more stable form than the enol form that it is in. The reformation of the ketone double bond is started by the deprotonation of that oxygen off the alpha carbon (C#2) by the same Lysine that protonated the oxygen in the first place.[7] The lone pair of electrons moves down kicking off the lone pairs that were making the double bond. This lone pair of electrons pulls a proton off the Tyrosine that deprotonated the carboxyl group in the decarboxylation step. The reason that we can say that the Lys and Tyr residues will be the same from the previous step because they are helping in holding the isocitrate molecule in the active site of the enzyme. These two residues will be able to hydrogen bond back and forth as long as they’re close enough to the substrate.[8]


Left: The isocitrate dehydrogenase enzyme as stated above produces alpha-ketoglutarate, carbon dioxide, and NADH + H+/NADPH + H+. There are three changes that occurred throughout the reaction. The oxidation of Carbon 2, the decarboxylation (loss of carbon dioxide) off Carbon 3, and the formation of a ketone group with a stereochemical change from sp3 to sp2.

Porcine Mitochondrial NADP+-dependent Isocitrate Dehydrogenase Complexed with Mn2+ and Isocitrate.[9]

Right: Surface view of the active site pocket where isocitrate is bounded by polar amino acids.[9]

Porcine Mitochondrial NADP+-dependent Isocitrate Dehydrogenase Complexed with Mn2+ and Isocitrate.[9]
Porcine Enzyme complex; Active site isocitrate and adjacent A.A.[9]

Above Left: The Porcine Heart Mitochondrial Enzyme.[9] In this picture you can see that the enzyme is a homeodimer where it is made up of two identical subunits and therefore two identical active sites. Above Right: This is the active site bared down to just isocitrate, Mn2+ and the surrounding interactive amino acids.[9]

Active site

The Isocitrate Dehydrogenase (IDH) enzyme structure in Escherichia coli was the first structure to be built and understood.[7] Since then, Escherichia coli has been used by most researchers to make comparisons to other isocitrate dehydrogenase enzymes. There is much detailed knowledge about this bacterial enzyme, and it has been found that most isocitrate dehydrogenases are similar in structure and therefore function. This similarity of structure and function gives reason to believe that the structures are conserved as well as the amino acids.[5] Therefore, the active sites amongst most prokaryotic isocitrate dehydrogenase enzymes should be conserved as well, which is observed throughout many studies done on prokaryotic enzymes. Eukaryotic isocitrate dehydrogenase enzymes on the other hand, have not been fully discovered yet. Each dimer of IDH has two active sites.[7] Each active site binds a NAD+/NADP+ molecule and a metal ion (Mg2+,Mn2+). In general, each active site has a conserved sequence of amino acids for each specific binding site. In Desulfotalea psychrophila (DpIDH)[7] and porcine (PcIDH)[9] there are three substrates bound to the active site.

(1) Isocitrate binds within the active site to a conserved sequence of about eight amino acids through hydrogen bonds. These acids include (may vary in residue but with similar properties) tyrosine, serine, asparagine, arginine, arginine, arginine, tyrosine, and lysine. Their positions on the backbone vary but they are all within a close range (i.e. Arg131 DpIDH and Arg133 PcIDH, Tyr138 DpIDH and Tyr140 PcIDH).[7]

(2) The metal ion (Mg2+, Mn2+) binds to three conserved amino acids through hydrogen bonds. These amino acids include three Aspartate residues.[7]

(3) NAD+ and NADP+ bind within the active site within four regions with similar properties amongst IDH enzymes. These regions vary but are around [250-260], [280-290], [300-330], and [365-380]. Again regions vary but the proximity of regions are conserved.[7]

Porcine IDH complex, Arg AA stabilizing isocitrate in the active site.[9]

Above: In this picture, residues Arg110, Arg133, and Arg101 are the three main stabilizing amino acids. They help to hold isocitrate in the active site and in the right orientation for isocitrate dehydrogenase to take place.[9]

Monomeric and dimeric IDH

When comparing monomer dehydrogenases with dimer dehydrogenases, it is important to note that most enzymes are dimers, to be specific, homodimers (two identical monomer subunits forming one dimeric unit). In a study done comparing Corynebacterium glutamicum and Escherichia coli[8], monomer and dimer, respectively, both enzymes were found to "efficiently catalyze identical reactions." However, C. glutamicum was recorded as having ten times as much activity than E. coli and seven times more affinitive/specific for NADP. C. glutamicum favored NADP+ over NAD+. In terms of stability with response to temperature, both enzymes had a similar Tm or melting temperature at about 55oC to 60oC. However, the monomer C. glutamicum showed a more consistent stability at higher temperatures, which was expected. The dimer E. coli showed stability at a higher temperature than normal due to the interactions between the two monomeric subunits.

One active site on the Porcine NADP+ dependent enzyme (green).[9]

Left: Porcine Heart Mitochondrial NADP+-dependent enzyme showing an active spot in green. Porcine enzyme is a homeodimer and has another active site on the other side.[9] Right: Escherichia coli was the first isocitrate dehydrogenase structure derived. There are three active sites. Three isocitrates, one isocitrate in the binding site for NADP+.

Clinical significance

Specific mutations in the isocitrate dehydrogenase gene IDH1 have been found in 12 percent of glioblastoma multiforme, with mutations found in nearly all cases of secondary glioblastomas, which develop from lower-grade gliomas, but rarely in primary high-grade glioblastoma multiforme. Patients whose tumor had an IDH1 mutation had longer survival. It is not known how the IDH1 mutation contributes to development of glioblastoma multiforme.[10]

See also


  1. ^ PDB 1CW7; Cherbavaz DB, Lee ME, Stroud RM, Koshland DE (January 2000). "Active site water molecules revealed in the 2.1 A resolution structure of a site-directed mutant of isocitrate dehydrogenase". J. Mol. Biol. 295 (3): 377–85. doi:10.1006/jmbi.1999.3195. PMID 10623532. 
  2. ^ Corpas FJ et al. (1999). "Peroxisomal NADP-Dependent Isocitrate Dehydrogenase. Characterization and Activity Regulation during Natural Senescence". Plant Physiology 121 (3): 921–928.. doi:10.1104/pp.121.3.921. PMID 10557241. 
  3. ^ a b c d e Tadhg P. Begley; McMurry, John (2005). The Organic Chemistry of Biological Pathways. Roberts and Co. Publishers. pp. 189-190. ISBN 0-9747077-1-6. 
  4. ^ a b c d e Cox, Michael; Nelson, David R.; Lehninger, Albert L (2005). Lehninger Principles of Biochemistry. San Francisco: W.H. Freeman. pp. 609-611. ISBN 0-7167-4339-6. 
  5. ^ a b Yasutake Y, Watanabe S, Yao M, Takada Y, Fukunaga N, Tanaka I (2003). "Crystal Structure of the Monomeric Isocitrate Dehydrogenase in the Presence of NADP+". Journal of Biological Chemistry 278 (38): 36897–36904. doi:10.1074/jbc.M304091200. PMID 12855708. 
  6. ^ Aoshima M, Igarashi Y (March 2008). "Nondecarboxylating and decarboxylating isocitrate dehydrogenases: oxalosuccinate reductase as an ancestral form of isocitrate dehydrogenase". Journal of bacteriology 190 (6): 2050–5. doi:10.1128/JB.01799-07. PMID 18203822. 
  7. ^ a b c d e f g h i j k l Fedøy AE, Yang N, Martinez A, Leiros HK, Steen IH (September 2007). "Structural and functional properties of isocitrate dehydrogenase from the psychrophilic bacterium Desulfotalea psychrophila reveal a cold-active enzyme with an unusual high thermal stability". J. Mol. Biol. 372 (1): 130–49. doi:10.1016/j.jmb.2007.06.040. PMID 17632124. 
  8. ^ a b Chen R, Yang H (November 2000). "A highly specific monomeric isocitrate dehydrogenase from Corynebacterium glutamicum". Arch. Biochem. Biophys. 383 (2): 238–45. doi:10.1006/abbi.2000.2082. PMID 11185559. 
  9. ^ a b c d e f g h i j k PDB 1LWD; Ceccarelli C, Neil B (2002). "The Crystal Structure of Porcine Mitochondrial NADP+-dependent Isocitrate Dehydrogenase Complexed with Mn2+ and Isocitrate". Journal of Biological Chemistry 277 (45): 43454–43462. doi:10.1074/jbc.M207306200. PMID 12207025. 
  10. ^ Parsons DW, Jones S, Zhang X, Lin JC, Leary RJ, Angenendt P, Mankoo P, Carter H, Siu IM, Gallia GL, Olivi A, McLendon R, Rasheed BA, Keir S, Nikolskaya T, Nikolsky Y, Busam DA, Tekleab H, Diaz LA, Hartigan J, Smith DR, Strausberg RL, Marie SK, Shinjo SM, Yan H, Riggins GJ, Bigner DD, Karchin R, Papadopoulos N, Parmigiani G, Vogelstein B, Velculescu VE, Kinzler KW (September 2008). "An integrated genomic analysis of human glioblastoma multiforme". Science 321 (5897): 1807–12. doi:10.1126/science.1164382. PMID 18772396. 


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