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Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB

[Image: see text] The process known as “adaptive response” allows Escherichia coli to respond to small doses of DNA-methylating agents by upregulating the expression of four proteins. While the role of three of these proteins in mitigating DNA damage is well understood, the function of AidB is less...

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Autores principales: Hamill, Michael J., Jost, Marco, Wong, Cintyu, Elliott, Sean J., Drennan, Catherine L.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Chemical Society 2011
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3217306/
https://www.ncbi.nlm.nih.gov/pubmed/22004173
http://dx.doi.org/10.1021/bi201340t
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author Hamill, Michael J.
Jost, Marco
Wong, Cintyu
Elliott, Sean J.
Drennan, Catherine L.
author_facet Hamill, Michael J.
Jost, Marco
Wong, Cintyu
Elliott, Sean J.
Drennan, Catherine L.
author_sort Hamill, Michael J.
collection PubMed
description [Image: see text] The process known as “adaptive response” allows Escherichia coli to respond to small doses of DNA-methylating agents by upregulating the expression of four proteins. While the role of three of these proteins in mitigating DNA damage is well understood, the function of AidB is less clear. Although AidB is a flavoprotein, no catalytic role has been established for the bound cofactor. Here we investigate the possibility that flavin plays a structural role in the assembly of the AidB tetramer. We report the generation and biophysical characterization of deflavinated AidB and of an AidB mutant that has greatly reduced affinity for flavin adenine dinucleotide (FAD). Using fluorescence quenching and analytical ultracentrifugation, we find that apo AidB has a high affinity for FAD, as indicated by an apparent dissociation constant of 402.1 ± 35.1 nM, and that binding of substoichiometric amounts of FAD triggers a transition in the AidB oligomeric state. In particular, deflavinated AidB is dimeric, whereas the addition of FAD yields a tetramer. We further investigate the dimerization and tetramerization interfaces of AidB by determining a 2.8 Å resolution crystal structure in space group P3(2) that contains three intact tetramers in the asymmetric unit. Taken together, our findings provide strong evidence that FAD plays a structural role in the formation of tetrameric AidB.
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spelling pubmed-32173062011-11-16 Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB Hamill, Michael J. Jost, Marco Wong, Cintyu Elliott, Sean J. Drennan, Catherine L. Biochemistry [Image: see text] The process known as “adaptive response” allows Escherichia coli to respond to small doses of DNA-methylating agents by upregulating the expression of four proteins. While the role of three of these proteins in mitigating DNA damage is well understood, the function of AidB is less clear. Although AidB is a flavoprotein, no catalytic role has been established for the bound cofactor. Here we investigate the possibility that flavin plays a structural role in the assembly of the AidB tetramer. We report the generation and biophysical characterization of deflavinated AidB and of an AidB mutant that has greatly reduced affinity for flavin adenine dinucleotide (FAD). Using fluorescence quenching and analytical ultracentrifugation, we find that apo AidB has a high affinity for FAD, as indicated by an apparent dissociation constant of 402.1 ± 35.1 nM, and that binding of substoichiometric amounts of FAD triggers a transition in the AidB oligomeric state. In particular, deflavinated AidB is dimeric, whereas the addition of FAD yields a tetramer. We further investigate the dimerization and tetramerization interfaces of AidB by determining a 2.8 Å resolution crystal structure in space group P3(2) that contains three intact tetramers in the asymmetric unit. Taken together, our findings provide strong evidence that FAD plays a structural role in the formation of tetrameric AidB. American Chemical Society 2011-10-17 2011-11-22 /pmc/articles/PMC3217306/ /pubmed/22004173 http://dx.doi.org/10.1021/bi201340t Text en Copyright © 2011 American Chemical Society http://pubs.acs.org This is an open-access article distributed under the ACS AuthorChoice Terms & Conditions. Any use of this article, must conform to the terms of that license which are available at http://pubs.acs.org.
spellingShingle Hamill, Michael J.
Jost, Marco
Wong, Cintyu
Elliott, Sean J.
Drennan, Catherine L.
Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB
title Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB
title_full Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB
title_fullStr Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB
title_full_unstemmed Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB
title_short Flavin-Induced Oligomerization in Escherichia coli Adaptive Response Protein AidB
title_sort flavin-induced oligomerization in escherichia coli adaptive response protein aidb
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3217306/
https://www.ncbi.nlm.nih.gov/pubmed/22004173
http://dx.doi.org/10.1021/bi201340t
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