An efficient NMR approach for obtaining sequence‐specific resonance assignments of larger proteins based on multiple isotopic labeling

Abstract
By simultaneously incorporating in a protein 13C‐carbonyl‐ and 15N‐labeled amino acids with different levels of enrichment, characteristic asymmetric doublet‐like patterns are observed for 15N nuclei that are directly adjacent to the 13C1‐labeled residues, providing unambiguous identification of a large number of unique dipeptide fragments of the protein. Additional assignments and qualitative structural information can be obtained from such a selectively labeled protein by recording multiple bond correlation spectra. The procedure is demonstrated for the protein calmodulin, complexed with calcium.