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Residue proximity information and protein model discrimination using saturation-suppressor mutagenesis

Sahoo, Anusmita and Khare, Shruti and Devanarayanan, Sivasankar and Jain, Pankaj C and Varadarajan, Raghavan (2016) Residue proximity information and protein model discrimination using saturation-suppressor mutagenesis. In: ELIFE, 4 .

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Official URL: http://dx.doi.org/10.7554/eLife.09532

Abstract

Identification of residue-residue contacts from primary sequence can be used to guide protein structure prediction. Using Escherichia coli CcdB as the test case, we describe an experimental method termed saturation-suppressor mutagenesis to acquire residue contact information. In this methodology, for each of five inactive CcdB mutants, exhaustive screens for suppressors were performed. Proximal suppressors were accurately discriminated from distal suppressors based on their phenotypes when present as single mutants. Experimentally identified putative proximal pairs formed spatial constraints to recover >98% of native-like models of CcdB from a decoy dataset. Suppressor methodology was also applied to the integral membrane protein, diacylglycerol kinase A where the structures determined by X-ray crystallography and NMR were significantly different. Suppressor as well as sequence co-variation data clearly point to the Xray structure being the functional one adopted in vivo. The methodology is applicable to any macromolecular system for which a convenient phenotypic assay exists.

Item Type: Journal Article
Additional Information: Copy right for this article belongs to the ELIFE SCIENCES PUBLICATIONS LTD, SHERATON HOUSE, CASTLE PARK, CAMBRIDGE, CB3 0AX, ENGLAND
Department/Centre: Division of Biological Sciences > Molecular Biophysics Unit
Depositing User: Id for Latest eprints
Date Deposited: 12 May 2016 07:34
Last Modified: 22 Nov 2018 15:04
URI: http://eprints.iisc.ac.in/id/eprint/53798

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