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Sci Rep. 2017 Feb 9;7:42343. doi: 10.1038/srep42343.

Structural variability of E. coli thioredoxin captured in the crystal structures of single-point mutants.

Author information

1
Universidad de Buenos Aires, Facultad de Farmacia y Bioquímica, Instituto de Química y Fisicoquímica Biológicas, CONICET, Junín 956, C1113AAD, Buenos Aires, Argentina.
2
Laboratorio de Fisicoquímica Biológica, Instituto de Química Biológica and Center for Free Radical and Biomedical Research, Universidad de la República, Montevideo, Uruguay.
3
Department of Integrative Biology, IGBMC, CNRS, INSERM, Université de Strasbourg, Illkirch, France.
4
Instituto de Biología Molecular y Celular de Rosario (IBR), Rosario, Santa Fe, Argentina.

Abstract

Thioredoxin is a ubiquitous small protein that catalyzes redox reactions of protein thiols. Additionally, thioredoxin from E. coli (EcTRX) is a widely-used model for structure-function studies. In a previous paper, we characterized several single-point mutants of the C-terminal helix (CTH) that alter global stability of EcTRX. However, spectroscopic signatures and enzymatic activity for some of these mutants were found essentially unaffected. A comprehensive structural characterization at the atomic level of these near-invariant mutants can provide detailed information about structural variability of EcTRX. We address this point through the determination of the crystal structures of four point-mutants, whose mutations occurs within or near the CTH, namely L94A, E101G, N106A and L107A. These structures are mostly unaffected compared with the wild-type variant. Notably, the E101G mutant presents a large region with two alternative traces for the backbone of the same chain. It represents a significant shift in backbone positions. Enzymatic activity measurements and conformational dynamics studies monitored by NMR and molecular dynamic simulations show that E101G mutation results in a small effect in the structural features of the protein. We hypothesize that these alternative conformations represent samples of the native-state ensemble of EcTRX, specifically the magnitude and location of conformational heterogeneity.

PMID:
28181556
PMCID:
PMC5299410
DOI:
10.1038/srep42343
[Indexed for MEDLINE]
Free PMC Article

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