6x9a: Difference between revisions
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==Structure of proline utilization A with trans-4-hydroxy-D-proline bound in the L-glutamate-gamma-semialdehyde dehydrogenase active site== | ==Structure of proline utilization A with trans-4-hydroxy-D-proline bound in the L-glutamate-gamma-semialdehyde dehydrogenase active site== | ||
<StructureSection load='6x9a' size='340' side='right'caption='[[6x9a]]' scene=''> | <StructureSection load='6x9a' size='340' side='right'caption='[[6x9a]], [[Resolution|resolution]] 1.41Å' scene=''> | ||
== Structural highlights == | == Structural highlights == | ||
<table><tr><td colspan='2'>Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6X9A OCA]. For a <b>guided tour on the structure components</b> use [http://proteopedia.org/fgij/fg.htm?mol=6X9A FirstGlance]. <br> | <table><tr><td colspan='2'>[[6x9a]] is a 2 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6X9A OCA]. For a <b>guided tour on the structure components</b> use [http://proteopedia.org/fgij/fg.htm?mol=6X9A FirstGlance]. <br> | ||
</td></tr><tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://proteopedia.org/fgij/fg.htm?mol=6x9a FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6x9a OCA], [http://pdbe.org/6x9a PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6x9a RCSB], [http://www.ebi.ac.uk/pdbsum/6x9a PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6x9a ProSAT]</span></td></tr> | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene>, <scene name='pdbligand=FMT:FORMIC+ACID'>FMT</scene>, <scene name='pdbligand=PEG:DI(HYDROXYETHYL)ETHER'>PEG</scene>, <scene name='pdbligand=PGE:TRIETHYLENE+GLYCOL'>PGE</scene>, <scene name='pdbligand=SO4:SULFATE+ION'>SO4</scene>, <scene name='pdbligand=UY7:(4S)-4-hydroxy-D-proline'>UY7</scene></td></tr> | ||
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://proteopedia.org/fgij/fg.htm?mol=6x9a FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6x9a OCA], [http://pdbe.org/6x9a PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6x9a RCSB], [http://www.ebi.ac.uk/pdbsum/6x9a PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6x9a ProSAT]</span></td></tr> | |||
</table> | </table> | ||
== Function == | |||
[[http://www.uniprot.org/uniprot/F7X6I3_SINMM F7X6I3_SINMM]] Oxidizes proline to glutamate for use as a carbon and nitrogen source.[PIRNR:PIRNR000197] | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Proline utilization A (PutA) proteins are bifunctional proline catabolic enzymes that catalyze the 4-electron oxidation of l-proline to l-glutamate using spatially-separated proline dehydrogenase and l-glutamate-gamma-semialdehyde dehydrogenase (GSALDH, a.k.a. ALDH4A1) active sites. The observation that l-proline inhibits both the GSALDH activity of PutA and monofunctional GSALDHs motivated us to study the inhibition of PutA by proline stereoisomers and analogs. Here we report five high-resolution crystal structures of PutA with the following ligands bound in the GSALDH active site: d-proline, trans-4-hydroxy-d-proline, cis-4-hydroxy-d-proline, l-proline, and trans-4-hydroxy-l-proline. Three of the structures are of ternary complexes of the enzyme with an inhibitor and either NAD(+) or NADH. To our knowledge, the NADH complex is the first for any GSALDH. The structures reveal a conserved mode of recognition of the inhibitor carboxylate, which results in the pyrrolidine rings of the d- and l-isomers having different orientations and different hydrogen bonding environments. Activity assays show that the compounds are weak inhibitors with millimolar inhibition constants. Curiously, although the inhibitors occupy the aldehyde binding site, kinetic measurements show the inhibition is uncompetitive. Uncompetitive inhibition may involve proline binding to a remote site or to the enzyme-NADH complex. Together, the structural and kinetic data expand our understanding of how proline-like molecules interact with GSALDH, reveal insight into the relationship between stereochemistry and inhibitor affinity, and demonstrate the pitfalls of inferring the mechanism of inhibition from crystal structures alone. | |||
Structural analysis of prolines and hydroxyprolines binding to the l-glutamate-gamma-semialdehyde dehydrogenase active site of bifunctional proline utilization A.,Campbell AC, Bogner AN, Mao Y, Becker DF, Tanner JJ Arch Biochem Biophys. 2020 Dec 18;698:108727. doi: 10.1016/j.abb.2020.108727. PMID:33333077<ref>PMID:33333077</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
<div class="pdbe-citations 6x9a" style="background-color:#fffaf0;"></div> | |||
== References == | |||
<references/> | |||
__TOC__ | __TOC__ | ||
</StructureSection> | </StructureSection> | ||
[[Category: Large Structures]] | [[Category: Large Structures]] | ||
[[Category: Campbell | [[Category: Campbell, A C]] | ||
[[Category: Tanner | [[Category: Tanner, J J]] | ||
[[Category: Aldehyde dehydrogenase]] | |||
[[Category: Bifunctional enzyme]] | |||
[[Category: Flavoenzyme]] | |||
[[Category: Oxidoreductase]] | |||
[[Category: Proline catabolism]] | |||
[[Category: Proline dehydrognease]] | |||
[[Category: Rossmann fold]] | |||
[[Category: Substrate channeling]] | |||
Revision as of 12:04, 6 January 2021
Structure of proline utilization A with trans-4-hydroxy-D-proline bound in the L-glutamate-gamma-semialdehyde dehydrogenase active siteStructure of proline utilization A with trans-4-hydroxy-D-proline bound in the L-glutamate-gamma-semialdehyde dehydrogenase active site
Structural highlights
Function[F7X6I3_SINMM] Oxidizes proline to glutamate for use as a carbon and nitrogen source.[PIRNR:PIRNR000197] Publication Abstract from PubMedProline utilization A (PutA) proteins are bifunctional proline catabolic enzymes that catalyze the 4-electron oxidation of l-proline to l-glutamate using spatially-separated proline dehydrogenase and l-glutamate-gamma-semialdehyde dehydrogenase (GSALDH, a.k.a. ALDH4A1) active sites. The observation that l-proline inhibits both the GSALDH activity of PutA and monofunctional GSALDHs motivated us to study the inhibition of PutA by proline stereoisomers and analogs. Here we report five high-resolution crystal structures of PutA with the following ligands bound in the GSALDH active site: d-proline, trans-4-hydroxy-d-proline, cis-4-hydroxy-d-proline, l-proline, and trans-4-hydroxy-l-proline. Three of the structures are of ternary complexes of the enzyme with an inhibitor and either NAD(+) or NADH. To our knowledge, the NADH complex is the first for any GSALDH. The structures reveal a conserved mode of recognition of the inhibitor carboxylate, which results in the pyrrolidine rings of the d- and l-isomers having different orientations and different hydrogen bonding environments. Activity assays show that the compounds are weak inhibitors with millimolar inhibition constants. Curiously, although the inhibitors occupy the aldehyde binding site, kinetic measurements show the inhibition is uncompetitive. Uncompetitive inhibition may involve proline binding to a remote site or to the enzyme-NADH complex. Together, the structural and kinetic data expand our understanding of how proline-like molecules interact with GSALDH, reveal insight into the relationship between stereochemistry and inhibitor affinity, and demonstrate the pitfalls of inferring the mechanism of inhibition from crystal structures alone. Structural analysis of prolines and hydroxyprolines binding to the l-glutamate-gamma-semialdehyde dehydrogenase active site of bifunctional proline utilization A.,Campbell AC, Bogner AN, Mao Y, Becker DF, Tanner JJ Arch Biochem Biophys. 2020 Dec 18;698:108727. doi: 10.1016/j.abb.2020.108727. PMID:33333077[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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