Cloning, expression, refolding, purification and preliminary crystallographic analysis of the sensory domain of the Campylobacter chemoreceptor for aspartate A (CcaA)

Mayra A. Machuca, Yu C. Liu, Anna Roujeinikova

Research output: Contribution to journalArticleResearchpeer-review

Abstract

In Campylobacter jejuni, chemotaxis and motility have been identified as important virulence factors that are required for host colonization and invasion. Chemotactic recognition of extracellular signals is mediated by the periplasmic sensory domains of its transducer-like proteins (Tlps). In this study, the sensory domain of the C. jejuni chemoreceptor for aspartate A (CcaA) has been expressed in Escherichia coli and purified from inclusion bodies. The urea-denatured protein was refolded and then crystallized by the hanging-drop vapour-diffusion method using PEG 3350 as a precipitating agent. A complete data set has been collected to 1.4 Å resolution using cryocooling conditions and synchrotron radiation. The crystals belonged to space group P1, with unit-cell parameters a = 39.3, b = 43.3, c = 50.9 Å, α = 92.5, β = 111.4, γ = 114.7°.
Original languageEnglish
Pages (from-to)110-113
Number of pages4
JournalActa Crystallographica. Section F: Structural Biology Communications
VolumeF71
Issue numberPart 1
DOIs
Publication statusPublished - 2015

Keywords

  • campylobacter jejuni
  • chemotaxis
  • transducer-like proteins
  • methyl-accepting proteins

Cite this

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title = "Cloning, expression, refolding, purification and preliminary crystallographic analysis of the sensory domain of the Campylobacter chemoreceptor for aspartate A (CcaA)",
abstract = "In Campylobacter jejuni, chemotaxis and motility have been identified as important virulence factors that are required for host colonization and invasion. Chemotactic recognition of extracellular signals is mediated by the periplasmic sensory domains of its transducer-like proteins (Tlps). In this study, the sensory domain of the C. jejuni chemoreceptor for aspartate A (CcaA) has been expressed in Escherichia coli and purified from inclusion bodies. The urea-denatured protein was refolded and then crystallized by the hanging-drop vapour-diffusion method using PEG 3350 as a precipitating agent. A complete data set has been collected to 1.4 {\AA} resolution using cryocooling conditions and synchrotron radiation. The crystals belonged to space group P1, with unit-cell parameters a = 39.3, b = 43.3, c = 50.9 {\AA}, α = 92.5, β = 111.4, γ = 114.7°.",
keywords = "campylobacter jejuni, chemotaxis, transducer-like proteins, methyl-accepting proteins",
author = "Machuca, {Mayra A.} and Liu, {Yu C.} and Anna Roujeinikova",
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T1 - Cloning, expression, refolding, purification and preliminary crystallographic analysis of the sensory domain of the Campylobacter chemoreceptor for aspartate A (CcaA)

AU - Machuca, Mayra A.

AU - Liu, Yu C.

AU - Roujeinikova, Anna

PY - 2015

Y1 - 2015

N2 - In Campylobacter jejuni, chemotaxis and motility have been identified as important virulence factors that are required for host colonization and invasion. Chemotactic recognition of extracellular signals is mediated by the periplasmic sensory domains of its transducer-like proteins (Tlps). In this study, the sensory domain of the C. jejuni chemoreceptor for aspartate A (CcaA) has been expressed in Escherichia coli and purified from inclusion bodies. The urea-denatured protein was refolded and then crystallized by the hanging-drop vapour-diffusion method using PEG 3350 as a precipitating agent. A complete data set has been collected to 1.4 Å resolution using cryocooling conditions and synchrotron radiation. The crystals belonged to space group P1, with unit-cell parameters a = 39.3, b = 43.3, c = 50.9 Å, α = 92.5, β = 111.4, γ = 114.7°.

AB - In Campylobacter jejuni, chemotaxis and motility have been identified as important virulence factors that are required for host colonization and invasion. Chemotactic recognition of extracellular signals is mediated by the periplasmic sensory domains of its transducer-like proteins (Tlps). In this study, the sensory domain of the C. jejuni chemoreceptor for aspartate A (CcaA) has been expressed in Escherichia coli and purified from inclusion bodies. The urea-denatured protein was refolded and then crystallized by the hanging-drop vapour-diffusion method using PEG 3350 as a precipitating agent. A complete data set has been collected to 1.4 Å resolution using cryocooling conditions and synchrotron radiation. The crystals belonged to space group P1, with unit-cell parameters a = 39.3, b = 43.3, c = 50.9 Å, α = 92.5, β = 111.4, γ = 114.7°.

KW - campylobacter jejuni

KW - chemotaxis

KW - transducer-like proteins

KW - methyl-accepting proteins

UR - http://www.ncbi.nlm.nih.gov/pubmed/25615981

U2 - 10.1107/S2053230X14027381

DO - 10.1107/S2053230X14027381

M3 - Article

VL - F71

SP - 110

EP - 113

JO - Acta Crystallographica. Section F: Structural Biology Communications

JF - Acta Crystallographica. Section F: Structural Biology Communications

SN - 1744-3091

IS - Part 1

ER -