Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.

We report the biochemical and biophysical characterization of outer membrane protein X (OmpX), an eight-stranded transmembrane β-barrel from E. coli, and compare the barrel behavior with a mutant devoid of methionine residues. Transmembrane outer membrane proteins of bacterial origin are known to di...

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Autores principales:	Deepti Chaturvedi, Radhakrishnan Mahalakshmi
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/e3261dd7bbc74cabac1153b5a9064e60
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spelling	oai:doaj.org-article:e3261dd7bbc74cabac1153b5a9064e602021-11-18T08:47:15ZMethionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.1932-620310.1371/journal.pone.0079351https://doaj.org/article/e3261dd7bbc74cabac1153b5a9064e602013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24265768/?tool=EBIhttps://doaj.org/toc/1932-6203We report the biochemical and biophysical characterization of outer membrane protein X (OmpX), an eight-stranded transmembrane β-barrel from E. coli, and compare the barrel behavior with a mutant devoid of methionine residues. Transmembrane outer membrane proteins of bacterial origin are known to display high tolerance to sequence rearrangements and mutations. Our studies with the triple mutant of OmpX that is devoid of all internal methionine residues (M18L; M21L; M118L) indicate that Met replacement has no influence on the refolding efficiency and structural characteristics of the protein. Surprisingly, the conserved substitution of Met→Leu leads to barrel destabilization and causes a lowering of the unfolding free energy by a factor of ∼8.5 kJ/mol, despite the mutations occurring at the loop regions. We report that the barrel destabilization is accompanied by a loss in cooperativity of unfolding in the presence of chemical denaturants. Furthermore, we are able to detect an unfolding intermediate in the Met-less barrel, whereas the parent protein exhibits a classic two-state unfolding. Thermal denaturation measurements also suggest a greater susceptibility of the OmpX barrel to heat, in the Met-less construct. Our studies reveal that even subtle variations in the extra-membrane region of rigid barrel structures such as OmpX, may bear severe implications on barrel stability. We propose that methionines contribute to efficient barrel structuring and protein-lipid interactions, and are therefore important elements of OmpX stability.Deepti ChaturvediRadhakrishnan MahalakshmiPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 11, p e79351 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Deepti Chaturvedi Radhakrishnan Mahalakshmi Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.
description	We report the biochemical and biophysical characterization of outer membrane protein X (OmpX), an eight-stranded transmembrane β-barrel from E. coli, and compare the barrel behavior with a mutant devoid of methionine residues. Transmembrane outer membrane proteins of bacterial origin are known to display high tolerance to sequence rearrangements and mutations. Our studies with the triple mutant of OmpX that is devoid of all internal methionine residues (M18L; M21L; M118L) indicate that Met replacement has no influence on the refolding efficiency and structural characteristics of the protein. Surprisingly, the conserved substitution of Met→Leu leads to barrel destabilization and causes a lowering of the unfolding free energy by a factor of ∼8.5 kJ/mol, despite the mutations occurring at the loop regions. We report that the barrel destabilization is accompanied by a loss in cooperativity of unfolding in the presence of chemical denaturants. Furthermore, we are able to detect an unfolding intermediate in the Met-less barrel, whereas the parent protein exhibits a classic two-state unfolding. Thermal denaturation measurements also suggest a greater susceptibility of the OmpX barrel to heat, in the Met-less construct. Our studies reveal that even subtle variations in the extra-membrane region of rigid barrel structures such as OmpX, may bear severe implications on barrel stability. We propose that methionines contribute to efficient barrel structuring and protein-lipid interactions, and are therefore important elements of OmpX stability.
format	article
author	Deepti Chaturvedi Radhakrishnan Mahalakshmi
author_facet	Deepti Chaturvedi Radhakrishnan Mahalakshmi
author_sort	Deepti Chaturvedi
title	Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.
title_short	Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.
title_full	Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.
title_fullStr	Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.
title_full_unstemmed	Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.
title_sort	methionine mutations of outer membrane protein x influence structural stability and beta-barrel unfolding.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/e3261dd7bbc74cabac1153b5a9064e60
work_keys_str_mv	AT deeptichaturvedi methioninemutationsofoutermembraneproteinxinfluencestructuralstabilityandbetabarrelunfolding AT radhakrishnanmahalakshmi methioninemutationsofoutermembraneproteinxinfluencestructuralstabilityandbetabarrelunfolding
_version_	1718421355647467520

Methionine mutations of outer membrane protein X influence structural stability and beta-barrel unfolding.

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