Rapid NMR Assignments of Proteins by Using Optimized Combinatorial Selective Unlabeling

Dubey, Abhinav and Kadumuri, Rajashekar Varma and Jaipuria, Garima and Vadrevu, Ramakrishna and Atreya, Hanudatta S (2016) Rapid NMR Assignments of Proteins by Using Optimized Combinatorial Selective Unlabeling. In: CHEMBIOCHEM, 17 (4). pp. 334-340.

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Abstract

A new approach for rapid resonance assignments in proteins based on amino acid selective unlabeling is presented. The method involves choosing a set of multiple amino acid types for selective unlabeling and identifying specific tripeptides surrounding the labeled residues from specific 2D NMR spectra in a combinatorial manner. The methodology directly yields sequence specific assignments, without requiring a contiguously stretch of amino acid residues to be linked, and is applicable to deuterated proteins. We show that a 2D N-15,H-1]HSQC spectrum with two 2D spectra can result in approximate to 50% assignments. The methodology was applied to two proteins: an intrinsically disordered protein (12kDa) and the 29kDa (268 residue) -subunit of Escherichia coli tryptophan synthase, which presents a challenging case with spectral overlaps and missing peaks. The method can augment existing approaches and will be useful for applications such as identifying active-site residues involved in ligand binding, phosphorylation, or protein-protein interactions, even prior to complete resonance assignments.

Item Type:	Journal Article
Publication:	CHEMBIOCHEM
Publisher:	WILEY-V C H VERLAG GMBH
Additional Information:	Copy right for this article belongs to the WILEY-V C H VERLAG GMBH, POSTFACH 101161, 69451 WEINHEIM, GERMANY
Keywords:	isotope scrambling; NMR spectroscopy; proteins; selective unlabeling assignments; sequence specific resonance; triple resonance experiments
Department/Centre:	Division of Chemical Sciences > NMR Research Centre (Formerly Sophisticated Instruments Facility) Others
Date Deposited:	02 Apr 2016 07:56
Last Modified:	02 Apr 2016 07:56
URI:	http://eprints.iisc.ac.in/id/eprint/53551

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