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Title: Crystal structures of green fluorescent protein with the unnatural amino acid 4-nitro-L -phenylalanine

Abstract

The X-ray crystal structures of two superfolder green fluorescent protein (sfGFP) constructs containing a genetically incorporated spectroscopic reporter unnatural amino acid, 4-nitro-L-phenylalanine (pNO 2F), at two unique sites in the protein have been determined. Amber codon-suppression methodology was used to site-specifically incorporate pNO 2F at a solvent-accessible (Asp133) and a partially buried (Asn149) site in sfGFP. The Asp133pNO 2F sfGFP construct crystallized with two molecules per asymmetric unit in space group P3 221 and the crystal structure was refined to 2.05 Å resolution. Crystals of Asn149pNO 2F sfGFP contained one molecule of sfGFP per asymmetric unit in space group P4 122 and the structure was refined to 1.60 Å resolution. The alignment of Asp133pNO 2F or Asn149pNO 2F sfGFP with wild-type sfGFP resulted in small root-mean-square deviations, illustrating that these residues do not significantly alter the protein structure and supporting the use of pNO 2F as an effective spectroscopic reporter of local protein structure and dynamics.

Authors:
; ; ; ; ORCiD logo
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
OTHER
OSTI Identifier:
1483086
Resource Type:
Journal Article
Journal Name:
Acta Crystallographica. Section F, Structural Biology Communications
Additional Journal Information:
Journal Volume: 74; Journal Issue: 10; Journal ID: ISSN 2053-230X
Publisher:
International Union of Crystallography
Country of Publication:
United States
Language:
ENGLISH

Citation Formats

Maurici, Nicole, Savidge, Nicole, Lee, Byung Uk, Brewer, Scott H., and Phillips-Piro, Christine M. Crystal structures of green fluorescent protein with the unnatural amino acid 4-nitro-L -phenylalanine. United States: N. p., 2018. Web. doi:10.1107/S2053230X1801169X.
Maurici, Nicole, Savidge, Nicole, Lee, Byung Uk, Brewer, Scott H., & Phillips-Piro, Christine M. Crystal structures of green fluorescent protein with the unnatural amino acid 4-nitro-L -phenylalanine. United States. doi:10.1107/S2053230X1801169X.
Maurici, Nicole, Savidge, Nicole, Lee, Byung Uk, Brewer, Scott H., and Phillips-Piro, Christine M. Wed . "Crystal structures of green fluorescent protein with the unnatural amino acid 4-nitro-L -phenylalanine". United States. doi:10.1107/S2053230X1801169X.
@article{osti_1483086,
title = {Crystal structures of green fluorescent protein with the unnatural amino acid 4-nitro-L -phenylalanine},
author = {Maurici, Nicole and Savidge, Nicole and Lee, Byung Uk and Brewer, Scott H. and Phillips-Piro, Christine M.},
abstractNote = {The X-ray crystal structures of two superfolder green fluorescent protein (sfGFP) constructs containing a genetically incorporated spectroscopic reporter unnatural amino acid, 4-nitro-L-phenylalanine (pNO2F), at two unique sites in the protein have been determined. Amber codon-suppression methodology was used to site-specifically incorporate pNO2F at a solvent-accessible (Asp133) and a partially buried (Asn149) site in sfGFP. The Asp133pNO2F sfGFP construct crystallized with two molecules per asymmetric unit in space group P3221 and the crystal structure was refined to 2.05 Å resolution. Crystals of Asn149pNO2F sfGFP contained one molecule of sfGFP per asymmetric unit in space group P4122 and the structure was refined to 1.60 Å resolution. The alignment of Asp133pNO2F or Asn149pNO2F sfGFP with wild-type sfGFP resulted in small root-mean-square deviations, illustrating that these residues do not significantly alter the protein structure and supporting the use of pNO2F as an effective spectroscopic reporter of local protein structure and dynamics.},
doi = {10.1107/S2053230X1801169X},
journal = {Acta Crystallographica. Section F, Structural Biology Communications},
issn = {2053-230X},
number = 10,
volume = 74,
place = {United States},
year = {2018},
month = {9}
}

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