PDBsum entry 4zf5

Fluorescent protein

PDB id: 4zf5

Contents

Protein chains

225 a.a.

Waters ×58

PDB id:

4zf5

Name:

Fluorescent protein

Title:

Crystal structure of green fluorescent protein (gfp); s65t, y66(cl2y), h148d; circular permutant ( 50-51)

Structure:

Green fluorescent protein. Chain: a, b. Engineered: yes. Mutation: yes

Source:

Aequorea victoria. Jellyfish. Organism_taxid: 6100. Gene: gfp. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

1.70Å R-factor: 0.199 R-free: 0.232

Authors:

L.M.Oltrogge,S.G.Boxer

Key ref:

L.M.Oltrogge and S.G.Boxer (2015). Short Hydrogen Bonds and Proton Delocalization in Green Fluorescent Protein (GFP). Acs Cent Sci, 1, 148-156. PubMed id: 27162964 DOI: 10.1021/acscentsci.5b00160

Date:

21-Apr-15 Release date: 10-Jun-15

Protein chains

P42212  (GFP_AEQVI) -  Green fluorescent protein from Aequorea victoria

Seq: 238 a.a.

Struc: 225 a.a.*

* PDB and UniProt seqs differ at 22 residue positions (black crosses)

DOI no: 10.1021/acscentsci.5b00160

Acs Cent Sci 1:148-156 (2015)

PubMed id: 27162964

Short Hydrogen Bonds and Proton Delocalization in Green Fluorescent Protein (GFP).

L.M.Oltrogge, S.G.Boxer.

ABSTRACT

Short hydrogen bonds and specifically low-barrier hydrogen bonds (LBHBs) have been the focus of much attention and controversy for their possible role in enzymatic catalysis. The green fluorescent protein (GFP) mutant S65T, H148D has been found to form a very short hydrogen bond between Asp148 and the chromophore resulting in significant spectral perturbations. Leveraging the unique autocatalytically formed chromophore and its sensitivity to this interaction we explore the consequences of proton affinity matching across this putative LBHB. Through the use of noncanonical amino acids introduced through nonsense suppression or global incorporation, we systematically modify the acidity of the GFP chromophore with halogen substituents. X-ray crystal structures indicated that the length of the interaction with Asp148 is unchanged at ∼2.45 Å while the absorbance spectra demonstrate an unprecedented degree of color tuning with increasing acidity. We utilized spectral isotope effects, isotope fractionation factors, and a simple 1D model of the hydrogen bond coordinate in order to gain insight into the potential energy surface and particularly the role that proton delocalization may play in this putative short hydrogen bond. The data and model suggest that even with the short donor-acceptor distance (∼2.45 Å) and near perfect affinity matching there is not a LBHB, that is, the barrier to proton transfer exceeds the H zero-point energy.