Expression, purification, crystallization and preliminary X-ray analysis of eCGP123, an extremely stable monomeric green fluorescent protein with reversible photoswitching properties

Craig Don Paul, Daouda Traore, Emma Byres, Jamie Rossjohn, Rodney Devenish, Csaba Kiss, Andrew Bradbury, Matthew Wilce, Mark Prescott

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Enhanced consensus green protein variant 123 (eCGP123) is an extremely thermostable green fluorescent protein (GFP) that exhibits useful negative reversible photoswitching properties. eCGP123 was derived by the application of both a consensus engineering approach and a recursive evolutionary process. Diffraction-quality crystals of recombinant eCGP123 were obtained by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant. The eCGP123 crystal diffracted X-rays to 2.10 A Es resolution. The data were indexed in space group P1, with unit-cell parameters a = 74.63, b = 75.38, c = 84.51 A Es , a = 90.96, b = 89.92, y = 104.03o. The Matthews coefficient (VM = 2.26 A Es 3 Da-1) and a solvent content of 46 indicated that the asymmetric unit contained eight eCGP123 molecules.
Original languageEnglish
Pages (from-to)1266 - 1268
Number of pages3
JournalActa Crystallographica. Section F: Structural Biology Communications
Volume67
Issue number10
DOIs
Publication statusPublished - 2011

Cite this

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title = "Expression, purification, crystallization and preliminary X-ray analysis of eCGP123, an extremely stable monomeric green fluorescent protein with reversible photoswitching properties",
abstract = "Enhanced consensus green protein variant 123 (eCGP123) is an extremely thermostable green fluorescent protein (GFP) that exhibits useful negative reversible photoswitching properties. eCGP123 was derived by the application of both a consensus engineering approach and a recursive evolutionary process. Diffraction-quality crystals of recombinant eCGP123 were obtained by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant. The eCGP123 crystal diffracted X-rays to 2.10 A Es resolution. The data were indexed in space group P1, with unit-cell parameters a = 74.63, b = 75.38, c = 84.51 A Es , a = 90.96, b = 89.92, y = 104.03o. The Matthews coefficient (VM = 2.26 A Es 3 Da-1) and a solvent content of 46 indicated that the asymmetric unit contained eight eCGP123 molecules.",
author = "{Don Paul}, Craig and Daouda Traore and Emma Byres and Jamie Rossjohn and Rodney Devenish and Csaba Kiss and Andrew Bradbury and Matthew Wilce and Mark Prescott",
year = "2011",
doi = "10.1107/S1744309111028156",
language = "English",
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pages = "1266 -- 1268",
journal = "Acta Crystallographica. Section F: Structural Biology Communications",
issn = "1744-3091",
publisher = "International Union of Crystallography",
number = "10",

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TY - JOUR

T1 - Expression, purification, crystallization and preliminary X-ray analysis of eCGP123, an extremely stable monomeric green fluorescent protein with reversible photoswitching properties

AU - Don Paul, Craig

AU - Traore, Daouda

AU - Byres, Emma

AU - Rossjohn, Jamie

AU - Devenish, Rodney

AU - Kiss, Csaba

AU - Bradbury, Andrew

AU - Wilce, Matthew

AU - Prescott, Mark

PY - 2011

Y1 - 2011

N2 - Enhanced consensus green protein variant 123 (eCGP123) is an extremely thermostable green fluorescent protein (GFP) that exhibits useful negative reversible photoswitching properties. eCGP123 was derived by the application of both a consensus engineering approach and a recursive evolutionary process. Diffraction-quality crystals of recombinant eCGP123 were obtained by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant. The eCGP123 crystal diffracted X-rays to 2.10 A Es resolution. The data were indexed in space group P1, with unit-cell parameters a = 74.63, b = 75.38, c = 84.51 A Es , a = 90.96, b = 89.92, y = 104.03o. The Matthews coefficient (VM = 2.26 A Es 3 Da-1) and a solvent content of 46 indicated that the asymmetric unit contained eight eCGP123 molecules.

AB - Enhanced consensus green protein variant 123 (eCGP123) is an extremely thermostable green fluorescent protein (GFP) that exhibits useful negative reversible photoswitching properties. eCGP123 was derived by the application of both a consensus engineering approach and a recursive evolutionary process. Diffraction-quality crystals of recombinant eCGP123 were obtained by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant. The eCGP123 crystal diffracted X-rays to 2.10 A Es resolution. The data were indexed in space group P1, with unit-cell parameters a = 74.63, b = 75.38, c = 84.51 A Es , a = 90.96, b = 89.92, y = 104.03o. The Matthews coefficient (VM = 2.26 A Es 3 Da-1) and a solvent content of 46 indicated that the asymmetric unit contained eight eCGP123 molecules.

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DO - 10.1107/S1744309111028156

M3 - Article

VL - 67

SP - 1266

EP - 1268

JO - Acta Crystallographica. Section F: Structural Biology Communications

JF - Acta Crystallographica. Section F: Structural Biology Communications

SN - 1744-3091

IS - 10

ER -