PDBsum entry 1q4c

Luminescent protein

PDB id: 1q4c

Contents

Protein chain

227 a.a. *

Waters ×231

* Residue conservation analysis

PDB id:

1q4c

Name:

Luminescent protein

Title:

S65t q80r t203c green fluorescent protein (gfp) ph 8.5

Structure:

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

Source:

Aequorea victoria. Organism_taxid: 6100. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Resolution:

1.55Å R-factor: 0.194 R-free: 0.214

Authors:

R.K.Jain,R.Ranganathan

Key ref:

R.K.Jain and R.Ranganathan (2004). Local complexity of amino acid interactions in a protein core. Proc Natl Acad Sci U S A, 101, 111-116. PubMed id: 14684834 DOI: 10.1073/pnas.2534352100

Date:

02-Aug-03 Release date: 03-Feb-04

Protein chain

P42212  (GFP_AEQVI) -  Green fluorescent protein from Aequorea victoria

Seq: 238 a.a.

Struc: 227 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

DOI no: 10.1073/pnas.2534352100

Proc Natl Acad Sci U S A 101:111-116 (2004)

PubMed id: 14684834

Local complexity of amino acid interactions in a protein core.

R.K.Jain, R.Ranganathan.

ABSTRACT

Atomic resolution structures of proteins indicate that the core is typically well packed, suggesting a densely connected network of interactions between amino acid residues. The combinatorial complexity of energetic interactions in such a network could be enormous, a problem that limits our ability to relate structure and function. Here, we report a case study of the complexity of amino acid interactions in a localized region within the core of the GFP, a particularly stable and tightly packed molecule. Mutations at three sites within the chromophore-binding pocket display an overlapping pattern of conformational change and are thermodynamically coupled, seemingly consistent with the dense network model. However, crystallographic and energetic analyses of coupling between mutations paint a different picture; pairs of mutations couple through independent "hotspots" in the region of structural overlap. The data indicate that, even in highly stable proteins, the core contains sufficient plasticity in packing to uncouple high-order energetic interactions of residues, a property that is likely general in proteins.

Selected figure(s)

Figure 1.
Fig. 1. Energetic characterization of the GFP chromophore-binding pocket. (a) Stereoview of the binding pocket viewed down the -barrel axis showing sites included in the mutagenic scan. The p-hydroxybenzylideneimidazolin-one chromophore is shown in green. (b) Mutagenic scan of the chromophore environment including the perturbation of pH shift from 8.5 to 5.5 ( pH). The energetic effect of each mutation is measured as change in chromophore absorbance maximum, a property that derives from changes to the ground state structure of GFP (24). Mutation of some sites has no significant energetic effect despite direct interaction with the chromophore (H148C), whereas the largest effect is seen for Q183, which only indirectly contacts the chromophore. This and subsequent figures were prepared by using GL-RENDER (L. Esser, personal communication), POVRAY (34), and RASTER3D (35).

Figure 4.
Fig. 4. Structure cycle analysis shows independent interaction mechanisms for the two-way thermodynamic couplings. Bar graphs (a and c) and colorimetric representations (b and d) of the magnitude of structural coupling ( r[norm]) for each atom in the T203C- pH (a and b) and Y145C-T203C (c and d) cycles. The values report the degree to which each atom feels the effect of one mutation differently when in the background of another mutation and is the structural analog of the double mutant cycle. Despite two-way thermodynamic coupling (Fig. 3) and overlapping structural change (Fig. 2) of the single mutants, the structural cycle analysis predicts that T203C and pH interact through a distinct mechanism from that of the T203C-Y145C pair.

Figures were selected by an automated process.