Fumarase 2
From Proteopedia
(Difference between revisions)
| Line 3: | Line 3: | ||
===Overview=== | ===Overview=== | ||
| - | Fumarase, also known as fumarate hydratase, is an enzyme in | + | Fumarase, also known as fumarate hydratase, is an enzyme in the citric acid cycle. In the seventh step of the reaction pathway, fumarase catalyzes the reversible hydration reaction that converts fumarate to malate and vice versa. |
===Stucture=== | ===Stucture=== | ||
| - | Fumarase is classified as an all alpha protein which belongs to the L-aspartase/fumarase family | + | Fumarase is classified as an all alpha protein which belongs to the L-aspartase/fumarase family. It forms a tetramer of identical subunits. Crystal structures of fumarase C revealed that the enzyme has two dicarboxylate binding sites; one was called the A site, and the second, the B site. This raises the question: which of the two sites is the active site of the enzyme? The A site shows relatively little change upon substrate binding, while the B site shifts substantially. <ref name="Weaver, et al."> Weaver,T. Structure of free fumarase C from ''Escherichia coli''. ''Acta Crystallographica'' (2005), '''D61''', 1395-1401. ['''http://dx.doi.org/10.1107/S0907444905024194''' doi:10.1107/S0907444905024194]</ref>. But these changes could account for regulation...so which site is the true active site? |
| + | In order to answer this question, an experiment that tested each of the sites independently was conducted. Both sites contain histidine residues: <scene name='72/726367/His_188/1'>His 188</scene> in the A-site and <scene name='72/726367/His_129/1'>His 129</scene> in the B-site. These sites were mutated to asparagine in separate experiments, and the effect on kinetics was measured. The results of the experiment showed that the H129N mutation had little effect on the enzymatic activity of the enzyme, as the specific activity of the enzyme was comparable to the wild-type enzyme. In contrast, the <scene name='72/726367/Ans_188_mutant/1'>H188N</scene> mutation dramatically reduced the specific activity of the catalytic reaction. These data strongly suggested that the H188 residue had a direct role in the catalytic mechanism of the enzyme and, therefore, that the H188 residue was located in the active site of the enzyme. This lead to the conclusion that that the A-site was in fact the active site of the enzyme<ref name= "Weaver">PMID:9098893</ref>. | ||
| - | == | + | == Structure and Function of the Fumurase Active Site == |
| - | The | + | The active site (A-site) of the fumarase enzyme is formed by residues from three of the enzyme’s four subunits and is located in a relatively deep pit that is removed from bulk solvent. The multi-subunit active site is comprised of atoms from residues 312-334 from subunit A, residues 182-200 from subunit C, and residues 129-145 from subunit D<ref>PMID: 7552727</ref>. The residues that form the active site are N141b, T100b, S98b, E331c, K324c, N326c, His 188C, and a water molecule, W-26. It is speculated that the H188 and W-26 are two of the most vital active site residues. Furthermore, H188 and W-26 form a short hydrogen bond, which increases the basicity of the water molecule. This electron-withdrawing hydrogen bond allows the water molecule to remove the C3 proton of <scene name='72/726367/L-malate/1'>L-malate</scene>. The cationic charge on W-26 plays an essential role in the stabilization of the double negative charge that is present on the aci-carboxylate at C4. Complex hydrogen bonding patterns in the active site involving T187, N141, H188, N362, and K324 also help stabilize the aci-carboxylate intermediate<ref name= "Weaver">PMID:9098893</ref>. By increasing the stabilization if the intermediate, the fumarase enzyme can effectively catalyze the hydration/dehydration reaction between L-malate and fumarate. |
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
| - | + | ||
===Regulation=== | ===Regulation=== | ||
| Line 24: | Line 20: | ||
| - | ===Other Interesting Information=== | ||
| - | Fumarase expression mainly occurs in skin, parathyroid, lymph, and colon tissues, and it is present throughout all life stages, from early development to mature adults. Fumarase comprises two specific classes which relate to the enzyme's: arrangement of subunits, metal ion requirement, and thermal stability. Class I fumarase isozymes can change their state, become inactive upon exposure to heat or radiation, are sensitive to superoxide anions, and Fe2+ dependent. Class II includes fumarase found in eukaryotes and prokaryotes, and they are iron-independent and thermally stable. | ||
| - | Mutations in the gene that encodes fumarase can lead to a deficiency in fumarase enzyme in the citric acid cycle which is known to cause certain diseases. Autosomal recessive mutants can result in fumarase deficiency, a metabolic disorder distinguished by excess fumarate in the body which can lead to severe developmental defects <ref>Remes, A., Rantala, H., Hiltunin, K., Leisti, J. & Ruokonen, A. Fumarase deficiency: two siblings with enlarged cerebral ventricles and polyhydramnios in utero. ''Pediatrics'' (1992), '''89'''(4), 730-734.</ref>. Inheritance of this autosomal recessive mutation has serious effects on early neural and brain development and can be fatal. Also, heterozygous fumarase mutations play a role in cancerous tumor development; specifically, the mutant H153R has identified as a factor in three families of malignant tumor growths <ref>Kokko, A., Ylisaukko-Oja, S., Kiuru, M., Takatalo, M., Salmikangas, P., Tuimala, J., et al. Modeling tumor predisposing FH mutations in yeast: effects on fumarase activity, growth phenotype and gene expression profile. ''Int. J. Cancer'' (2006), '''118'''(6), 1340-1345.</ref>. | ||
</structure section> | </structure section> | ||
===References=== | ===References=== | ||
<references/> | <references/> | ||
Revision as of 20:47, 8 February 2018
| |||||||||||
