PDBsum entry 1b9c

Luminescent protein

PDB id: 1b9c

Contents

Protein chains

225 a.a. *

Waters ×286

* Residue conservation analysis

PDB id:

1b9c

Name:

Luminescent protein

Title:

Green fluorescent protein mutant f99s, m153t and v163a

Structure:

Protein (green fluorescent protein). Chain: a, b, c, d. Engineered: yes. Mutation: yes

Source:

Aequorea victoria. Organism_taxid: 6100. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Other_details: the n-terminal his-tag has been removed

Biol. unit:

Tetramer (from PQS)

Resolution:

2.40Å R-factor: 0.204 R-free: 0.280

Authors:

R.Battistutta,A.Negro,G.Zanotti

Key ref:

R.Battistutta et al. (2000). Crystal structure and refolding properties of the mutant F99S/M153T/V163A of the green fluorescent protein. Proteins, 41, 429-437. PubMed id: 11056031 DOI: 10.1002/1097-0134(20001201)41:4<429::AID-PROT10>3.0.CO;2-D

Date:

09-Feb-99 Release date: 17-Nov-00

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 4 residue positions (black crosses)

DOI no: 10.1002/1097-0134(20001201)41:4<429::AID-PROT10>3.0.CO;2-D

Proteins 41:429-437 (2000)

PubMed id: 11056031

Crystal structure and refolding properties of the mutant F99S/M153T/V163A of the green fluorescent protein.

R.Battistutta, A.Negro, G.Zanotti.

ABSTRACT

The mutant F99S/M153T/V163A of the Green Fluorescent Protein (c3-GFP) has spectral characteristics similar to the wild-type GFP, but it is 42-fold more fluorescent in vivo. Here, we report the crystal structure and the refolding properties of c3-GFP and compare them with those of the less fluorescent wt-GFP and S65T mutant. The topology and the overall structure of c3-GFP is similar to the wild-type GFP. The three mutated residues, Ser99, Thr153, and Ala163, lie on the surface of the protein in three different beta-strands. The side chains of Ser99 and Thr153 are exposed to the solvent, whereas that of Ala163 points toward the interior of the protein. No significant deviation from the structure of the wild-type molecule is found around these positions, and there is not clear evidence of any distortion in the position of the chromophore or of the surrounding residues induced by the mutated amino acids. In vitro refolding experiments on urea-denatured c3-GFP reveal a renaturation behavior similar to that of the S65T molecule, with kinetic constants of the same order of magnitude. We conclude that the higher fluorescence activity of c3-GFP can be attributed neither to particular structural features nor to a faster folding process, as previously proposed.

Selected figure(s)

Figure 2.
Figure 2. Schematic drawing of the Green Fluorescent Protein (c3-GFP) dimer, viewed along the twofold axis. The chromophore (in gray) and the three mutated residues (in black) are represented as ball-and-stick.

Figure 3.
Figure 3. Stereo view of the 2Fo-Fc electron density map contoured at 1 around the chromophore (CRO) and the main interacting residues His148 (H148), Thr203 (T203), Ser205 (S205), and Glu222 (E222). Water molecules around the chromophore are not shown.

The above figures are reprinted by permission from John Wiley & Sons, Inc.: Proteins (2000, 41, 429-437) copyright 2000.

Figures were selected by an automated process.