Efficient sequential assignments in proteins with reduced dimensionality 3D HN(CA)NH |
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Authors: | Kousik Chandra Garima Jaipuria Divya Shet Hanudatta S Atreya |
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Institution: | (1) NMR Research Centre, Indian Institute of Science, Bangalore, 560012, India;(2) Solid State and Structural Chemistry Unit, Indian Institute of Science, Bangalore, 560012, India; |
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Abstract: | We present reduced dimensionality (RD) 3D HN(CA)NH for efficient sequential assignment in proteins. The experiment correlates the 15N and 1H chemical shift of a residue (‘i’) with those of its immediate N-terminal (i − 1) and C-terminal (i + 1) neighbors and provides four-dimensional chemical shift correlations rapidly with high resolution. An assignment strategy
is presented which combines the correlations observed in this experiment with amino acid type information obtained from 3D
CBCA(CO)NH. By classifying the 20 amino acid types into seven distinct categories based on 13Cβ chemical shifts, it is observed that a stretch of five sequentially connected residues is sufficient to map uniquely on to
the polypeptide for sequence specific resonance assignments. This method is exemplified by application to three different
systems: maltose binding protein (42 kDa), intrinsically disordered domain of insulin-like growth factor binding protein-2
and Ubiquitin. Fast data acquisition is demonstrated using longitudinal 1H relaxation optimization. Overall, 3D HN(CA)NH is a powerful tool for high throughput resonance assignment, in particular for unfolded or intrinsically disordered
polypeptides. |
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