Triose Phosphate Isomerase: Difference between revisions
Jump to navigation
Jump to search
Michal Harel (talk | contribs) No edit summary |
Michal Harel (talk | contribs) No edit summary |
||
| (20 intermediate revisions by 3 users not shown) | |||
| Line 1: | Line 1: | ||
{{BAMBED | |||
<StructureSection load='2ypi' size=' | |DATE=June 30, 2011 | ||
caption='TPI (yeast) at 2.5 Å [[resolution]] ([[2ypi]]) dimer. The two identical chains are | |OLDID=1265234 | ||
|BAMBEDDOI=10.1002/bmb.20550 | |||
}} | |||
<StructureSection load='2ypi' size='350' | |||
caption='TPI (yeast) at 2.5 Å [[resolution]] ([[2ypi]]) dimer. The two identical chains are in grey and green. Ligand is the inhibitor 2-phosphoglycolic acid (PGA). ' | |||
scene='' > | scene='' > | ||
[[Image:TriosePhosphateIsomerase_Ribbon_pastel_photo_small.jpg|thumb|left|260px| Ribbon drawing of the "TIM barrel" fold]] | [[Image:TriosePhosphateIsomerase_Ribbon_pastel_photo_small.jpg|thumb|left|260px| Ribbon drawing of the "TIM barrel" fold]] | ||
[[Triose Phosphate Isomerase]] (TPI or TIM) is a ubiquitous dimeric enzyme with a molecular weight of ~54 kD (27 kD per subunit) which catalyzes the reversible interconversion of the triose phosphate isomers <scene name='Triose_Phosphate_Isomerase/Morph/1'>dihydroxyacetone phosphate (DHAP)</scene> and <scene name='Triose_Phosphate_Isomerase/Morph/2'>D-glyceraldehyde-3-phosphate (GAP)</scene>, an essential process in the glycolytic pathway. More simply, the enzyme catalyzes the <scene name='Triose_Phosphate_Isomerase/Morph/3'>isomerization of a ketose (DHAP) to an aldose (GAP)</scene>, also referred to as '''PGAL'''. In regards to the two isomers, at equilibrium, roughly 96% of the triose phosphate is in the DHAP isomer form; however, the isomerization reaction proceeds due to the rapid removal of GAP from the subsequent reactions of glycolysis. The TPI tertiary structure is the classic example of the "TIM barrel" fold (see image at left). The TPI structure is shown on the right (PDB entry [[2ypi]]) in complex with the inhibitor 2-phosphoglycolic acid (PGA), which is bound to each of its two active sites. TPI is an example of a catalytically perfect enzyme, indicating that for almost every enzyme-substrate encounter, a product is formed and that this interaction is limited only by the substrate diffusion rate. | ==Introduction== | ||
[[Triose Phosphate Isomerase]] (TPI or TIM) is a ubiquitous dimeric enzyme with a molecular weight of ~54 kD (27 kD per subunit) which catalyzes the reversible interconversion of the triose phosphate isomers <scene name='Triose_Phosphate_Isomerase/Morph/1'>dihydroxyacetone phosphate (DHAP)</scene> and <scene name='Triose_Phosphate_Isomerase/Morph/2'>D-glyceraldehyde-3-phosphate (GAP)</scene>, an essential process in the glycolytic pathway. More simply, the enzyme catalyzes the <scene name='Triose_Phosphate_Isomerase/Morph/3'>isomerization of a ketose (DHAP) to an aldose (GAP)</scene>, also referred to as '''PGAL'''. In regards to the two isomers, at equilibrium, roughly 96% of the triose phosphate is in the DHAP isomer form; however, the isomerization reaction proceeds due to the rapid removal of GAP from the subsequent reactions of glycolysis. The TPI tertiary structure is the classic example of the "TIM barrel" fold (see image at left). The TPI structure is shown on the right (PDB entry [[2ypi]]) in complex with the inhibitor 2-phosphoglycolic acid (PGA), which is bound to each of its two active sites. TPI is an example of a catalytically perfect enzyme, indicating that for almost every enzyme-substrate encounter, a product is formed and that this interaction is limited only by the substrate diffusion rate. See [[Glycolysis Enzymes]] and [[Isomerases]]. | |||
In addition to its role in glycolysis, TPI is also involved in several additional metabolic biological processes including gluconeogenesis, the pentose phosphate shunt, and fatty acid biosynthesis. A point mutation to a glutamate residue (Glu104) of TPI results in triose phosphate isomerase deficiency, an autosomal recessive inherited disorder characterized by an increased accumulation of DHAP in erythrocytes. Structurally, this point mutation abolishes TPI’s ability to dimerize, subsequently inhibiting its catalytic activity. More details in [[ Triose Phosphate Isomerase Structure & Mechanism]] | In addition to its role in glycolysis, TPI is also involved in several additional metabolic biological processes including gluconeogenesis, the pentose phosphate shunt, and fatty acid biosynthesis. A point mutation to a glutamate residue (Glu104) of TPI results in triose phosphate isomerase deficiency, an autosomal recessive inherited disorder characterized by an increased accumulation of DHAP in erythrocytes. Structurally, this point mutation abolishes TPI’s ability to dimerize, subsequently inhibiting its catalytic activity. More details in<br /> | ||
* [[ Triose Phosphate Isomerase Structure & Mechanism]]<br /> | |||
* [[Derivation of Triose Phosphate Isomerase]]<br /> | |||
* [[Triosephosphate Isomerase]]. | |||
== Mechanism == | == Mechanism == | ||
| Line 47: | Line 55: | ||
{{Clear}} | {{Clear}} | ||
More recently a series of NMR experiments carried out by Mildvan and co-workers have shed light onto an alternative "Criss-cross" mechanism involving a LBHB between the catalytic Glu165 and the O1 oxygen of the substrate. This mechanism stipulates the His95 side chain does not directly transfer protons, this rather being accomplished entirely by Glu165. Support for this mechanism was provided by Richard and coworkers who carried out tritium labeling experiments demonstrating a significant amount of intramolecular transfer (49%) of the <sup>3</sup>H label from substrate (DHAP) to product (GAP)<ref>PMID:15709774</ref>. Using phosphoglycolohydroxamate (PGH), a mimic of the enediol(ate) intermediate, a 14.9 ppm chemical shift (6 ppm downfield) as well as a deuterium fractionation factor of 0.38 was observed with the TIM-PGH complex, corresponding to a highly deshielded proton involved in a LBHB between Glu165 and the hydroxamate oxygen of PGH. Conversely, the same NMR study found an additional hydrogen bond between the N-ε proton of His95 and the carbonyl oxygen of PGH; however, its chemical shift of 13.5 (0.4 ppm downfield from free enzyme) and fractionation factor of 0.71 indicated this was a strong H-bond, but not a LBHB.<ref>PMID:9748211</ref>. | More recently a series of NMR experiments carried out by Mildvan and co-workers have shed light onto an alternative "Criss-cross" mechanism involving a LBHB between the catalytic Glu165 and the O1 oxygen of the substrate. This mechanism stipulates the His95 side chain does not directly transfer protons, this rather being accomplished entirely by Glu165. Support for this mechanism was provided by Richard and coworkers who carried out tritium labeling experiments demonstrating a significant amount of intramolecular transfer (49%) of the <sup>3</sup>H label from substrate (DHAP) to product (GAP)<ref>PMID:15709774</ref>. Using phosphoglycolohydroxamate (PGH), a mimic of the enediol(ate) intermediate, a 14.9 ppm chemical shift (6 ppm downfield) as well as a deuterium fractionation factor of 0.38 was observed with the TIM-PGH complex, corresponding to a highly deshielded proton involved in a LBHB between Glu165 and the hydroxamate oxygen of PGH. Conversely, the same NMR study found an additional hydrogen bond between the N-ε proton of His95 and the carbonyl oxygen of PGH; however, its chemical shift of 13.5 (0.4 ppm downfield from free enzyme) and fractionation factor of 0.71 indicated this was a strong H-bond, but not a LBHB.<ref>PMID:9748211</ref>. | ||
[[Image: | [[Image:LBHB2_Glu1.png|left|thumb|500x250px|'''LBHB between Glu165 and DHAP''']] The formation of the LBHB between Glu165 and O1 of the inhibitor PGH is due to the matching of p''K''as and the alternative mechanism suggests that Glu-165, in addition to its role in initially abstracting the proton from the substrate, may also shuttle protons to and from the oxygens in the intermediate. Also, the "criss-cross" mechanism implies that, by donating a normal hydrogen bond, the role of His95 is to polarize the carbonyl oxygen and lower the p''K''a of PGH in order to facilitate subsequent proton abstraction<ref>PMID:9748211</ref>. It has been argued that that the LBHB formed between Glu165 and PGH is a consequence of using the inhibitor PGH, whose hydroxamate p''K''a of 9 better matches Glu165 then His95, and that the biological reaction would instead see the enediol forming a LHBH with His95, as mentioned above. Overall, the mechanism employed by TPI has yet to be completely solved and recent NMR studies involving both WT and mutant TPI enzymes have revealed contributions from both the "classic" and "criss-cross" mechanisms. | ||
[[Image:crisscross2.png|right|thumb|750x350px| ''' Alternative "Criss-Cross" TPI Mechanism Involving LBHB Between Glu165 and O1 of the Intermediate''']] | [[Image:crisscross2.png|right|thumb|750x350px| ''' Alternative "Criss-Cross" TPI Mechanism Involving LBHB Between Glu165 and O1 of the Intermediate''']] | ||
===Inhibitors of Triose Phosphate Isomerase=== | ===Inhibitors of Triose Phosphate Isomerase=== | ||
{{Clear}} | |||
[[Image:TPIinhibitors.png|thumb|left|400px| '''Inhibitors of Triose Phosphate Isomerase''']] | [[Image:TPIinhibitors.png|thumb|left|400px| '''Inhibitors of Triose Phosphate Isomerase''']] | ||
| Line 91: | Line 101: | ||
One specific example of sequence homology is that of loop 6 and loop 7 residues, whose structural contributions are discussed above. In a sequence alignment of 133 TIM sequences, two highly conserved motifs are noticed. First, 114 sequences in loop 6 contain the PXW sequence family (where X is I,L or V in 112 sequences or otherwise a T or K). Secondly, loop 7 contains a highly conserved YGGS motif; however, this motif is only found when the N-terminal hinge contains tryptophan. | One specific example of sequence homology is that of loop 6 and loop 7 residues, whose structural contributions are discussed above. In a sequence alignment of 133 TIM sequences, two highly conserved motifs are noticed. First, 114 sequences in loop 6 contain the PXW sequence family (where X is I,L or V in 112 sequences or otherwise a T or K). Secondly, loop 7 contains a highly conserved YGGS motif; however, this motif is only found when the N-terminal hinge contains tryptophan. | ||
== 3D Structures of triose phosphate isomerase== | == 3D Structures of triose phosphate isomerase== | ||
[[Triose phosphate isomerase 3D structures]] | |||
</StructureSection> | |||
== Additional Resources == | == Additional Resources == | ||
| Line 184: | Line 118: | ||
[[Category: Topic Page]] | [[Category: Topic Page]] | ||
[[Category:Featured in BAMBED]] | |||
<references/> | <references/> | ||