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Crystal structures of green fluorescent protein with the unnatural amino acid 4‐nitro‐ l ‐phenylalanine
Author(s) -
Maurici Nicole,
Savidge Nicole,
Lee Byung Uk,
Brewer Scott H.,
Phillips-Piro Christine M.
Publication year - 2018
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x1801169x
Subject(s) - chemistry , fluorescence , nitro , fluorescent protein , green fluorescent protein , phenylalanine , amino acid , molecule , stereochemistry , biochemistry , organic chemistry , gene , physics , quantum mechanics , alkyl
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 2 F), at two unique sites in the protein have been determined. Amber codon‐suppression methodology was used to site‐specifically incorporate pNO 2 F at a solvent‐accessible (Asp133) and a partially buried (Asn149) site in sfGFP. The Asp133pNO 2 F sfGFP construct crystallized with two molecules per asymmetric unit in space group P 3 2 21 and the crystal structure was refined to 2.05 Å resolution. Crystals of Asn149pNO 2 F sfGFP contained one molecule of sfGFP per asymmetric unit in space group P 4 1 22 and the structure was refined to 1.60 Å resolution. The alignment of Asp133pNO 2 F or Asn149pNO 2 F 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 2 F as an effective spectroscopic reporter of local protein structure and dynamics.