6own: Difference between revisions
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The | ==Crystal structure of Msm DnaB1 intein splicing domain bound with zinc== | ||
<StructureSection load='6own' size='340' side='right'caption='[[6own]], [[Resolution|resolution]] 1.95Å' scene=''> | |||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[6own]] is a 3 chain structure with sequence from [https://en.wikipedia.org/wiki/Mycolicibacterium_smegmatis Mycolicibacterium smegmatis]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6OWN OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=6OWN 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]] 1.95Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=ZN:ZINC+ION'>ZN</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=6own FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6own OCA], [https://pdbe.org/6own PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=6own RCSB], [https://www.ebi.ac.uk/pdbsum/6own PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=6own ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/A0R7F4_MYCS2 A0R7F4_MYCS2] Participates in initiation and elongation during chromosome replication; it exhibits DNA-dependent ATPase activity.[ARBA:ARBA00002835][RuleBase:RU362085] | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Inteins, as posttranslational regulatory elements, can tune protein function to environmental changes by conditional protein splicing (CPS). Translated as subdomains interrupting host proteins, inteins splice to scarlessly join flanking sequences (exteins). We used DnaB-intein1 (DnaBi1) from a replicative helicase of Mycobacterium smegmatis to build a kanamycin intein splicing reporter (KISR) that links splicing of DnaBi1 to kanamycin resistance. Using expression in heterologous Escherichia coli, we observed phenotypic classes of various levels of splicing-dependent resistance (SDR) and related these to the insertion position of DnaBi1 within the kanamycin resistance protein (KanR). The KanR-DnaBi1 construct demonstrating the most stringent SDR was used to probe for CPS of DnaB in the native host environment, M. smegmatis We show here that zinc, important during mycobacterial pathogenesis, inhibits DnaB splicing in M. smegmatis Using an in vitro reporter system, we demonstrated that zinc potently and reversibly inhibited DnaBi1 splicing, as well as splicing of a comparable intein from Mycobacterium leprae Finally, in a 1.95 A crystal structure, we show that zinc inhibits splicing through binding to the very cysteine that initiates the splicing reaction. Together, our results provide compelling support for a model whereby mycobacterial DnaB protein splicing, and thus DNA replication, is responsive to environmental zinc.IMPORTANCE Inteins are present in a large fraction of prokaryotes and localize within conserved proteins, including the mycobacterial replicative helicase DnaB. In addition to their extensive protein engineering applications, inteins have emerged as environmentally responsive posttranslational regulators of the genes that encode them. While several studies have shown compelling evidence of conditional protein splicing (CPS), examination of splicing in the native host of the intein has proven to be challenging. Here, we demonstrated through a number of measures, including the use of a splicing-dependent sensor capable of monitoring intein activity in the native host, that zinc is a potent and reversible inhibitor of mycobacterial DnaB splicing. This work also expands our knowledge of site selection for intein insertion within nonnative proteins, demonstrating that splicing-dependent host protein activation correlates with proximity to the active site. Additionally, we surmise that splicing regulation by zinc has mycobacteriocidal and CPS application potential. | |||
Conditional DnaB Protein Splicing Is Reversibly Inhibited by Zinc in Mycobacteria.,Woods D, Vangaveti S, Egbanum I, Sweeney AM, Li Z, Bacot-Davis V, LeSassier DS, Stanger M, Hardison GE, Li H, Belfort M, Lennon CW mBio. 2020 Jul 14;11(4). pii: mBio.01403-20. doi: 10.1128/mBio.01403-20. PMID:32665276<ref>PMID:32665276</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
[[Category: | </div> | ||
[[Category: | <div class="pdbe-citations 6own" style="background-color:#fffaf0;"></div> | ||
[[Category: Bacot-Davis | |||
==See Also== | |||
*[[Helicase 3D structures|Helicase 3D structures]] | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
[[Category: Large Structures]] | |||
[[Category: Mycolicibacterium smegmatis]] | |||
[[Category: Bacot-Davis VR]] | |||
[[Category: Belfort M]] | |||
[[Category: Lennon C]] | |||