PDBsum entry 2jt3

Structural protein

PDB id: 2jt3

Contents

Protein chain

161 a.a. *

* Residue conservation analysis

PDB id:

2jt3

Name:

Structural protein

Title:

Solution structure of f153w cardiac troponin c

Structure:

Troponin c. Chain: a. Synonym: tn-c. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: tnnc1, tnnc. Expressed in: escherichia coli. Expression_system_taxid: 562. Expression_system_variant: (de3)plyss.

NMR struc:

10 models

Authors:

X.Wang,P.Mercier,P.Letourneau,B.D.Sykes

Key ref:

X.Wang et al. (2005). Effects of Phe-to-Trp mutation and fluorotryptophan incorporation on the solution structure of cardiac troponin C, and analysis of its suitability as a potential probe for in situ NMR studies. Protein Sci, 14, 2447-2460. PubMed id: 16131667 DOI: 10.1110/ps.051595805

Date:

18-Jul-07 Release date: 31-Jul-07

Protein chain

P63316  (TNNC1_HUMAN) -  Troponin C, slow skeletal and cardiac muscles from Homo sapiens

Seq: 161 a.a.

Struc: 161 a.a.*

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

DOI no: 10.1110/ps.051595805

Protein Sci 14:2447-2460 (2005)

PubMed id: 16131667

Effects of Phe-to-Trp mutation and fluorotryptophan incorporation on the solution structure of cardiac troponin C, and analysis of its suitability as a potential probe for in situ NMR studies.

X.Wang, P.Mercier, P.J.Letourneau, B.D.Sykes.

ABSTRACT

19F NMR spectroscopy is potentially a powerful tool for probing protein properties in situ. However, results obtained using this technique are relevant only if the 19F probe offers minimal perturbation to the surrounding environment. In this paper, we examine the effect of 5-fluorotryptophan (5fW) incorporation on the three-dimensional structure of cardiac troponin-C (cTnC), with the intention of developing a 19F-labeled TnC for use in in situ 19FNMR. We find that, in general, 5fW does not perturb the structure of the protein significantly. Replacement of residue Phe 153 with 5fW produces no noticeable change in protein conformation. However, replacement of residue Phe 104 with 5fW produces a folding behavior that is dependent on the Escherichia coli strain used to express the mutant. The orientations of the indole rings in these mutants are such that the Trp residue adopts a chi2 of approximately 90 degrees in the F104W mutant and approximately -100 degrees in the F153W mutant. Using results from 19F-1H heteronuclear NOE experiment, we show the replacement of L-Trp with 5fW at these positions does not change the orientation of the indole ring and the spread of the 5fW side-chain dihedral angles increases moderately for the F104(5fW) mutant and not at all for the F153(5fW) mutant. Based on these structures, we conclude that the substitution of Phe by 5fW at these two positions has minimal effects on the structure of cTnC and that the 5fW indole rings in both mutants have well defined orientation, making the two mutants viable candidates for use in in situ 19F NMR spectroscopy.

Selected figure(s)

Figure 1.
Figure 1. (A) Ribbon representation of the C-terminal domain of the wild-type chicken cTnC. Side chains of F104 and F153 (sites of mutation) are labeled red and blue, respectively. (B) Indole ring of Tryptophan. Numbering of the carbon is illustrated. Each proton is labeled using nomenclature used in the paper. In 5fW, H 3 is replaced with a ^19F atom.

Figure 6.
Figure 6. Ribbon representations of backbone superimposition of the wild-type chicken cTnC structure (PDB code 1AJ4 [PDB] ) with (A) the F104W structure (red), (B) the F104(5fW) structure (gray), (C) the F153W structure (orange), (D) the F153(5fW) structure (yellow). The wild-type structure is shown in blue in all superimpositions. RMSDs between the mutant structures and the wild-type structure are shown below each ribbon representation. In all panels, the superimposition is done using residues in structured regions only (residues 95–103, 111–123, 130–140, and 147–158).

The above figures are reprinted by permission from the Protein Society: Protein Sci (2005, 14, 2447-2460) copyright 2005.

Figures were selected by an automated process.