View source for 8fo0


==The structure of a crystallizable variant of E. coli pyruvate formate-lyase activating enzyme bound to a partially cleaved SAM molecule==
<StructureSection load='8fo0' size='340' side='right'caption='[[8fo0]], [[Resolution|resolution]] 1.69&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[8fo0]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Escherichia_coli_K-12 Escherichia coli K-12]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=8FO0 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=8FO0 FirstGlance]. <br>
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.69&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=K:POTASSIUM+ION'>K</scene>, <scene name='pdbligand=SAM:S-ADENOSYLMETHIONINE'>SAM</scene></td></tr>
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=8fo0 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=8fo0 OCA], [https://pdbe.org/8fo0 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=8fo0 RCSB], [https://www.ebi.ac.uk/pdbsum/8fo0 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=8fo0 ProSAT]</span></td></tr>
</table>
== Function ==
[https://www.uniprot.org/uniprot/PFLA_ECOLI PFLA_ECOLI] Activation of pyruvate formate-lyase 1 under anaerobic conditions by generation of an organic free radical, using S-adenosylmethionine and reduced flavodoxin as cosubstrates to produce 5'-deoxy-adenosine.
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
Radical S-adenosyl-l-methionine (SAM) enzymes are ubiquitous in nature and carry out a broad variety of difficult chemical transformations initiated by hydrogen atom abstraction. Although numerous radical SAM (RS) enzymes have been structurally characterized, many prove recalcitrant to crystallization needed for atomic-level structure determination using X-ray crystallography, and even those that have been crystallized for an initial study can be difficult to recrystallize for further structural work. We present here a method for computationally engineering previously observed crystallographic contacts and employ it to obtain more reproducible crystallization of the RS enzyme pyruvate formate-lyase activating enzyme (PFL-AE). We show that the computationally engineered variant binds a typical RS [4Fe-4S](2+/+) cluster that binds SAM, with electron paramagnetic resonance properties indistinguishable from the native PFL-AE. The variant also retains the typical PFL-AE catalytic activity, as evidenced by the characteristic glycyl radical electron paramagnetic resonance signal observed upon incubation of the PFL-AE variant with reducing agent, SAM, and PFL. The PFL-AE variant was also crystallized in the [4Fe-4S](2+) state with SAM bound, providing a new high-resolution structure of the SAM complex in the absence of substrate. Finally, by incubating such a crystal in a solution of sodium dithionite, the reductive cleavage of SAM is triggered, providing us with a structure in which the SAM cleavage products 5'-deoxyadenosine and methionine are bound in the active site. We propose that the methods described herein may be useful in the structural characterization of other difficult-to-resolve proteins.

Computational engineering of previously crystallized pyruvate formate-lyase activating enzyme reveals insights into SAM binding and reductive cleavage.,Moody JD, Hill S, Lundahl MN, Saxton AJ, Galambas A, Broderick WE, Lawrence CM, Broderick JB J Biol Chem. 2023 Jun;299(6):104791. doi: 10.1016/j.jbc.2023.104791. Epub 2023 , May 6. PMID:37156396<ref>PMID:37156396</ref>

From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
</div>
<div class="pdbe-citations 8fo0" style="background-color:#fffaf0;"></div>
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Escherichia coli K-12]]
[[Category: Large Structures]]
[[Category: Broderick JB]]
[[Category: Galambas A]]
[[Category: Lawrence CM]]
[[Category: Moody JD]]
[[Category: Saxton AJ]]