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==R2-like ligand-binding oxidase A171F mutant with anaerobically reconstituted Mn/Fe cofactor== | ==R2-like ligand-binding oxidase A171F mutant with anaerobically reconstituted Mn/Fe cofactor== | ||
<StructureSection load='6f6b' size='340' side='right' caption='[[6f6b]], [[Resolution|resolution]] 2.01Å' scene=''> | <StructureSection load='6f6b' size='340' side='right'caption='[[6f6b]], [[Resolution|resolution]] 2.01Å' scene=''> | ||
== Structural highlights == | == Structural highlights == | ||
<table><tr><td colspan='2'>[[6f6b]] is a 1 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6F6B OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6F6B FirstGlance]. <br> | <table><tr><td colspan='2'>[[6f6b]] is a 1 chain structure with sequence from [http://en.wikipedia.org/wiki/Geoka Geoka]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6F6B OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6F6B FirstGlance]. <br> | ||
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=FE2:FE+(II)+ION'>FE2</scene>, <scene name='pdbligand=MN:MANGANESE+(II)+ION'>MN</scene></td></tr> | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=FE2:FE+(II)+ION'>FE2</scene>, <scene name='pdbligand=MN:MANGANESE+(II)+ION'>MN</scene></td></tr> | ||
<tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4hr0|4hr0]], [[4hr4|4hr4]], [[4hr5|4hr5]], [[4xb9|4xb9]], [[4xbv|4xbv]], [[4xbw|4xbw]], [[5dco|5dco]], [[5dcr|5dcr]], [[5dcs|5dcs]], [[5ekb|5ekb]], [[5omk|5omk]], [[5omj|5omj]], [[6f65|6f65]], [[6f6c|6f6c]], [[6f6e|6f6e]], [[6f6f|6f6f]], [[6f6g|6f6g]], [[6f6h|6f6h]], [[6f6k|6f6k]], [[6f6l|6f6l]], [[6f6m|6f6m]]</td></tr> | <tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4hr0|4hr0]], [[4hr4|4hr4]], [[4hr5|4hr5]], [[4xb9|4xb9]], [[4xbv|4xbv]], [[4xbw|4xbw]], [[5dco|5dco]], [[5dcr|5dcr]], [[5dcs|5dcs]], [[5ekb|5ekb]], [[5omk|5omk]], [[5omj|5omj]], [[6f65|6f65]], [[6f6c|6f6c]], [[6f6e|6f6e]], [[6f6f|6f6f]], [[6f6g|6f6g]], [[6f6h|6f6h]], [[6f6k|6f6k]], [[6f6l|6f6l]], [[6f6m|6f6m]]</td></tr> | ||
<tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">GK2771 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=235909 GEOKA])</td></tr> | |||
<tr id='activity'><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/Ribonucleoside-diphosphate_reductase Ribonucleoside-diphosphate reductase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.17.4.1 1.17.4.1] </span></td></tr> | <tr id='activity'><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/Ribonucleoside-diphosphate_reductase Ribonucleoside-diphosphate reductase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.17.4.1 1.17.4.1] </span></td></tr> | ||
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6f6b FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6f6b OCA], [http://pdbe.org/6f6b PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6f6b RCSB], [http://www.ebi.ac.uk/pdbsum/6f6b PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6f6b ProSAT]</span></td></tr> | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6f6b FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6f6b OCA], [http://pdbe.org/6f6b PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6f6b RCSB], [http://www.ebi.ac.uk/pdbsum/6f6b PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6f6b ProSAT]</span></td></tr> | ||
</table> | </table> | ||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
R2-like ligand-binding oxidases (R2lox) assemble a heterodinuclear Mn/Fe cofactor which performs reductive dioxygen (O2) activation, catalyzes formation of a tyrosine-valine ether cross-link in the protein scaffold, and binds a fatty acid in a putative substrate channel. We have previously shown that the N-terminal metal binding site 1 is unspecific for manganese or iron in the absence of O2, but prefers manganese in the presence of O2, whereas the C-terminal site 2 is specific for iron. Here, we analyze the effects of amino acid exchanges in the cofactor environment on cofactor assembly and metalation specificity using X-ray crystallography, X-ray absorption spectroscopy, and metal quantification. We find that exchange of either the cross-linking tyrosine or the valine, regardless of whether the mutation still allows cross-link formation or not, results in unspecific manganese or iron binding at site 1 both in the absence or presence of O2, while site 2 still prefers iron as in the wild-type. In contrast, a mutation that blocks binding of the fatty acid does not affect the metal specificity of either site under anoxic or aerobic conditions, and cross-link formation is still observed. All variants assemble a dinuclear trivalent metal cofactor in the aerobic resting state, independently of cross-link formation. These findings imply that the cross-link residues are required to achieve the preference for manganese in site 1 in the presence of O2. The metalation specificity, therefore, appears to be established during the redox reactions leading to cross-link formation. | |||
Assembly of a heterodinuclear Mn/Fe cofactor is coupled to tyrosine-valine ether cross-link formation in the R2-like ligand-binding oxidase.,Griese JJ, Kositzki R, Haumann M, Hogbom M J Biol Inorg Chem. 2019 Mar;24(2):211-221. doi: 10.1007/s00775-019-01639-4. Epub , 2019 Jan 28. PMID:30689052<ref>PMID:30689052</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
<div class="pdbe-citations 6f6b" style="background-color:#fffaf0;"></div> | |||
== References == | |||
<references/> | |||
__TOC__ | __TOC__ | ||
</StructureSection> | </StructureSection> | ||
[[Category: Geoka]] | |||
[[Category: Large Structures]] | |||
[[Category: Ribonucleoside-diphosphate reductase]] | [[Category: Ribonucleoside-diphosphate reductase]] | ||
[[Category: Griese, J J]] | [[Category: Griese, J J]] | ||
Latest revision as of 15:38, 13 March 2019
R2-like ligand-binding oxidase A171F mutant with anaerobically reconstituted Mn/Fe cofactorR2-like ligand-binding oxidase A171F mutant with anaerobically reconstituted Mn/Fe cofactor
Structural highlights
Publication Abstract from PubMedR2-like ligand-binding oxidases (R2lox) assemble a heterodinuclear Mn/Fe cofactor which performs reductive dioxygen (O2) activation, catalyzes formation of a tyrosine-valine ether cross-link in the protein scaffold, and binds a fatty acid in a putative substrate channel. We have previously shown that the N-terminal metal binding site 1 is unspecific for manganese or iron in the absence of O2, but prefers manganese in the presence of O2, whereas the C-terminal site 2 is specific for iron. Here, we analyze the effects of amino acid exchanges in the cofactor environment on cofactor assembly and metalation specificity using X-ray crystallography, X-ray absorption spectroscopy, and metal quantification. We find that exchange of either the cross-linking tyrosine or the valine, regardless of whether the mutation still allows cross-link formation or not, results in unspecific manganese or iron binding at site 1 both in the absence or presence of O2, while site 2 still prefers iron as in the wild-type. In contrast, a mutation that blocks binding of the fatty acid does not affect the metal specificity of either site under anoxic or aerobic conditions, and cross-link formation is still observed. All variants assemble a dinuclear trivalent metal cofactor in the aerobic resting state, independently of cross-link formation. These findings imply that the cross-link residues are required to achieve the preference for manganese in site 1 in the presence of O2. The metalation specificity, therefore, appears to be established during the redox reactions leading to cross-link formation. Assembly of a heterodinuclear Mn/Fe cofactor is coupled to tyrosine-valine ether cross-link formation in the R2-like ligand-binding oxidase.,Griese JJ, Kositzki R, Haumann M, Hogbom M J Biol Inorg Chem. 2019 Mar;24(2):211-221. doi: 10.1007/s00775-019-01639-4. Epub , 2019 Jan 28. PMID:30689052[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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