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The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode

The two-domain protein PduO, involved in 1,2-propanediol utilization in the pathogenic Gram-negative bacterium Salmonella enterica is an ATP:Cob(I)alamin adenosyltransferase, but this is a function of the N-terminal domain alone. The role of its C-terminal domain (PduOC) is, however, unknown. In thi...

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Autores principales: Ortiz de Orué Lucana, Darío, Hickey, Neal, Hensel, Michael, Klare, Johann P., Geremia, Silvano, Tiufiakova, Tatiana, Torda, Andrew E.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4923194/
https://www.ncbi.nlm.nih.gov/pubmed/27446048
http://dx.doi.org/10.3389/fmicb.2016.01010
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author Ortiz de Orué Lucana, Darío
Hickey, Neal
Hensel, Michael
Klare, Johann P.
Geremia, Silvano
Tiufiakova, Tatiana
Torda, Andrew E.
author_facet Ortiz de Orué Lucana, Darío
Hickey, Neal
Hensel, Michael
Klare, Johann P.
Geremia, Silvano
Tiufiakova, Tatiana
Torda, Andrew E.
author_sort Ortiz de Orué Lucana, Darío
collection PubMed
description The two-domain protein PduO, involved in 1,2-propanediol utilization in the pathogenic Gram-negative bacterium Salmonella enterica is an ATP:Cob(I)alamin adenosyltransferase, but this is a function of the N-terminal domain alone. The role of its C-terminal domain (PduOC) is, however, unknown. In this study, comparative growth assays with a set of Salmonella mutant strains showed that this domain is necessary for effective in vivo catabolism of 1,2-propanediol. It was also shown that isolated, recombinantly-expressed PduOC binds heme in vivo. The structure of PduOC co-crystallized with heme was solved (1.9 Å resolution) showing an octameric assembly with four heme moieities. The four heme groups are highly solvent-exposed and the heme iron is hexa-coordinated with bis-His ligation by histidines from different monomers. Static light scattering confirmed the octameric assembly in solution, but a mutation of the heme-coordinating histidine caused dissociation into dimers. Isothermal titration calorimetry using the PduOC apoprotein showed strong heme binding (K(d) = 1.6 × 10(−7) M). Biochemical experiments showed that the absence of the C-terminal domain in PduO did not affect adenosyltransferase activity in vitro. The evidence suggests that PduOC:heme plays an important role in the set of cobalamin transformations required for effective catabolism of 1,2-propanediol. Salmonella PduO is one of the rare proteins which binds the redox-active metabolites heme and cobalamin, and the heme-binding mode of the C-terminal domain differs from that in other members of this protein family.
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spelling pubmed-49231942016-07-21 The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode Ortiz de Orué Lucana, Darío Hickey, Neal Hensel, Michael Klare, Johann P. Geremia, Silvano Tiufiakova, Tatiana Torda, Andrew E. Front Microbiol Microbiology The two-domain protein PduO, involved in 1,2-propanediol utilization in the pathogenic Gram-negative bacterium Salmonella enterica is an ATP:Cob(I)alamin adenosyltransferase, but this is a function of the N-terminal domain alone. The role of its C-terminal domain (PduOC) is, however, unknown. In this study, comparative growth assays with a set of Salmonella mutant strains showed that this domain is necessary for effective in vivo catabolism of 1,2-propanediol. It was also shown that isolated, recombinantly-expressed PduOC binds heme in vivo. The structure of PduOC co-crystallized with heme was solved (1.9 Å resolution) showing an octameric assembly with four heme moieities. The four heme groups are highly solvent-exposed and the heme iron is hexa-coordinated with bis-His ligation by histidines from different monomers. Static light scattering confirmed the octameric assembly in solution, but a mutation of the heme-coordinating histidine caused dissociation into dimers. Isothermal titration calorimetry using the PduOC apoprotein showed strong heme binding (K(d) = 1.6 × 10(−7) M). Biochemical experiments showed that the absence of the C-terminal domain in PduO did not affect adenosyltransferase activity in vitro. The evidence suggests that PduOC:heme plays an important role in the set of cobalamin transformations required for effective catabolism of 1,2-propanediol. Salmonella PduO is one of the rare proteins which binds the redox-active metabolites heme and cobalamin, and the heme-binding mode of the C-terminal domain differs from that in other members of this protein family. Frontiers Media S.A. 2016-06-28 /pmc/articles/PMC4923194/ /pubmed/27446048 http://dx.doi.org/10.3389/fmicb.2016.01010 Text en Copyright © 2016 Ortiz de Orué Lucana, Hickey, Hensel, Klare, Geremia, Tiufiakova and Torda. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Ortiz de Orué Lucana, Darío
Hickey, Neal
Hensel, Michael
Klare, Johann P.
Geremia, Silvano
Tiufiakova, Tatiana
Torda, Andrew E.
The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode
title The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode
title_full The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode
title_fullStr The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode
title_full_unstemmed The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode
title_short The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode
title_sort crystal structure of the c-terminal domain of the salmonella enterica pduo protein: an old fold with a new heme-binding mode
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4923194/
https://www.ncbi.nlm.nih.gov/pubmed/27446048
http://dx.doi.org/10.3389/fmicb.2016.01010
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