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Caught in action: selecting peptide aptamers against intrinsically disordered proteins in live cells.
- Source :
-
Scientific reports [Sci Rep] 2015 Mar 24; Vol. 5, pp. 9402. Date of Electronic Publication: 2015 Mar 24. - Publication Year :
- 2015
-
Abstract
- Intrinsically disordered proteins (IDPs) or unstructured segments within proteins play an important role in cellular physiology and pathology. Low cellular concentration, multiple binding partners, frequent post-translational modifications and the presence of multiple conformations make it difficult to characterize IDP interactions in intact cells. We used peptide aptamers selected by using the yeast-two-hybrid scheme and in-cell NMR to identify high affinity binders to transiently structured IDP and unstructured segments at atomic resolution. Since both the selection and characterization of peptide aptamers take place inside the cell, only physiologically relevant conformations of IDPs are targeted. The method is validated by using peptide aptamers selected against the prokaryotic ubiquitin-like protein, Pup, of the mycobacterium proteasome. The selected aptamers bind to distinct sites on Pup and have vastly different effects on rescuing mycobacterial proteasome substrate and on the survival of the Bacille-Calmette-Guèrin, BCG, strain of M. bovis. This technology can be applied to study the elusive action of IDPs under near physiological conditions.
- Subjects :
- Amino Acid Sequence
Aptamers, Peptide pharmacology
Bacterial Proteins metabolism
Binding Sites
Intrinsically Disordered Proteins metabolism
Microbial Viability drug effects
Models, Molecular
Molecular Sequence Data
Mycobacterium bovis chemistry
Mycobacterium bovis drug effects
Mycobacterium bovis metabolism
Mycobacterium tuberculosis chemistry
Mycobacterium tuberculosis drug effects
Mycobacterium tuberculosis metabolism
Proteasome Endopeptidase Complex drug effects
Proteasome Endopeptidase Complex metabolism
Protein Binding
Protein Folding
Protein Structure, Secondary
Protein Structure, Tertiary
Two-Hybrid System Techniques
Ubiquitins metabolism
Aptamers, Peptide chemistry
Bacterial Proteins chemistry
Intrinsically Disordered Proteins chemistry
Proteasome Endopeptidase Complex chemistry
Protein Processing, Post-Translational
Ubiquitins chemistry
Subjects
Details
- Language :
- English
- ISSN :
- 2045-2322
- Volume :
- 5
- Database :
- MEDLINE
- Journal :
- Scientific reports
- Publication Type :
- Academic Journal
- Accession number :
- 25801767
- Full Text :
- https://doi.org/10.1038/srep09402