scholarly journals Inside Back Cover: Significant Expansion of the Fluorescent Protein Chromophore through the Genetic Incorporation of a Metal-Chelating Unnatural Amino Acid (Angew. Chem. Int. Ed. 18/2013)

2013 ◽  
Vol 52 (18) ◽  
pp. 4955-4955 ◽  
Author(s):  
Xiaohong Liu ◽  
Jiasong Li ◽  
Cheng Hu ◽  
Qing Zhou ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Fluorescent Protein ◽  
Unnatural Amino Acid ◽  
Back Cover ◽  
Metal Chelating ◽  
Genetic Incorporation

Significant Expansion of the Fluorescent Protein Chromophore through the Genetic Incorporation of a Metal-Chelating Unnatural Amino Acid

2013 ◽  
Vol 125 (18) ◽  
pp. 4905-4909 ◽  
Author(s):  
Xiaohong Liu ◽  
Jiasong Li ◽  
Cheng Hu ◽  
Qing Zhou ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Fluorescent Protein ◽  
Unnatural Amino Acid ◽  
Metal Chelating ◽  
Genetic Incorporation

Genetic Incorporation of a Metal-Chelating Amino Acid as a Probe for Protein Electron Transfer

2012 ◽  
Vol 124 (41) ◽  
pp. 10407-10411 ◽  
Author(s):  
Xiaohong Liu ◽  
Jiasong Li ◽  
Jianshu Dong ◽  
Cheng Hu ◽  
Weimin Gong ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Amino Acid ◽  
Metal Chelating ◽  
Protein Electron Transfer ◽  
Genetic Incorporation

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

2018 ◽  
Vol 74 (10) ◽  
pp. 650-655
Author(s):  
Nicole Maurici ◽  
Nicole Savidge ◽  
Byung Uk Lee ◽  
Scott H. Brewer ◽  
Christine M. Phillips-Piro
Keyword(s):  
Green Fluorescent Protein ◽  
Protein Structure ◽  
Amino Acid ◽  
Crystal Structures ◽  
Fluorescent Protein ◽  
Unnatural Amino Acid ◽  
Asymmetric Unit ◽  
Space Group ◽  
Green Fluorescent ◽  
Amber Codon Suppression

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.

Structural and spectrophotometric investigation of two unnatural amino-acid altered chromophores in the superfolder green fluorescent protein

2021 ◽  
Vol 77 (8) ◽  
Author(s):  
Gregory M. Olenginski ◽  
Juliana Piacentini ◽  
Darcy R. Harris ◽  
Nicolette A. Runko ◽  
Brianna M. Papoutsis ◽  
...  
Keyword(s):  
Amino Acids ◽  
Green Fluorescent Protein ◽  
Amino Acid ◽  
Unnatural Amino Acids ◽  
Fluorescent Protein ◽  
Photophysical Properties ◽  
Unnatural Amino Acid ◽  
Green Fluorescent ◽  
Absorbance Band ◽  
Protein Constructs

The spectrophotometric properties of the green fluorescent protein (GFP) result from the post-translationally cyclized chromophore composed of three amino acids including a tyrosine at the center of the β-barrel protein. Altering the amino acids in the chromophore or the nearby region has resulted in numerous GFP variants with differing photophysical properties. To further examine the effect of small atomic changes in the chromophore on the structure and photophysical properties of GFP, the hydroxyl group of the chromophore tyrosine was replaced with a nitro or a cyano group. The structures and spectrophotometric properties of these superfolder GFP (sfGFP) variants with the unnatural amino acids (UAAs) 4-nitro-L-phenylalanine or 4-cyano-L-phenylalanine were explored. Notably, the characteristic 487 nm absorbance band of wild-type (wt) sfGFP is absent in both unnatural amino-acid-containing protein constructs (Tyr66pNO2Phe-sfGFP and Tyr66pCNPhe-sfGFP). Consequently, neither Tyr66pNO2Phe-sfGFP nor Tyr66pCNPhe-sfGFP exhibited the characteristic emission of wt sfGFP centered at 511 nm when excited at 487 nm. Tyr66pNO2Phe-sfGFP appeared orange due to an absorbance band centered at 406 nm that was not present in wt sfGFP, while Tyr66pCNPhe-sfGFP appeared colorless with an absorbance band centered at 365 nm. Mass spectrometry and X-ray crystallography confirmed the presence of a fully formed chromophore and no significant structural changes in either of these UAA-containing protein constructs, signaling that the change in the observed photophysical properties of the proteins is the result of the presence of the UAA in the chromophore.

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