Isotopologues of a Metabolic Precursor for Selective N-15 and C-13 Histidine Labeling

Histidine is a versatile residue with distinct properties ensuring many proteins' structure and proper function. Its imidazole side-chain represents an ideal chemical entity to serve as a proton shuttle in enzyme mechanisms, control recognition interfaces either by contribution of its aromatic Pi system or in its cationic form, and acts as a coordinating ligand to metal cations. These functional capabilities are modulated by the local molecular environment, which influences pK_a values and tautomeric states. NMR spectroscopy has proven to be a reliable method for probing the distinct functions of histidine. Here, we describe the synthesis of isotopically labeled variants of a non-chiral precursor to introduce NMR active ¹³C and/or ¹⁵N nuclei into histidine side-chains. The compounds were employed to selectively label protein targets of significant interest in current drug discovery programs, such as the WD repeat containing protein 5 (WDR5), the Src homology 2 domain of the phospholipase C (PLCγ-SH2) and the product of the Kirsten rat sarcoma virus oncogene (KRAS).