Selective observation of the disordered import signal of a globular protein by in-cell NMR: the example of frataxins

Protein Science : a Publication of the Protein Society
Matija PopovicAnnalisa Pastore

Abstract

We have exploited the capability of in-cell NMR to selectively observe flexible regions within folded proteins to carry out a comparative study of two members of the highly conserved frataxin family which are found both in prokaryotes and in eukaryotes. They all contain a globular domain which shares more than 50% identity, which in eukaryotes is preceded by an N-terminal tail containing the mitochondrial import signal. We demonstrate that the NMR spectrum of the bacterial ortholog CyaY cannot be observed in the homologous E. coli system, although it becomes fully observable as soon as the cells are lysed. This behavior has been observed for several other compact globular proteins as seems to be the rule rather than the exception. The NMR spectrum of the yeast ortholog Yfh1 contains instead visible signals from the protein. We demonstrate that they correspond to the flexible N-terminal tail indicating that this is flexible and unfolded. This flexibility of the N-terminus agrees with previous studies of human frataxin, despite the extensive sequence diversity of this region in the two proteins. Interestingly, the residues that we observe in in-cell experiments are not visible in the crystal structure of a Yfh1 mutant designed to...Continue Reading

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Citations

May 16, 2017·Archives of Biochemistry and Biophysics·Eric B GibbsScott A Showalter
Aug 2, 2018·Cellular and Molecular Life Sciences : CMLS·Alexander V FoninVladimir N Uversky
Aug 11, 2021·The Biochemical Journal·Vladimir N Uversky
Aug 24, 2017·Biochemistry·Ciara Kyne, Peter B Crowley

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Methods Mentioned

BETA
nuclear magnetic resonance
NMR

Software Mentioned

NMRPipe
Biopack
STINT

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