Acireductone dioxygenase (ARD) is a 179-residue enzyme containing a paramagnetic Ni+2 ion in the active site. Because of electron-nuclear spin interactions, H-1 resonances within -9 angstrom of the Ni+2 are broadened beyond detection. For this reason, H-1-detected multidimensional NMR experiments are not suitable for structural characterization of the active site of ARD, and no isostructural diamagnetic homologue is available. Rapid recycle two-dimensional direct C-13 detection NMR methods previously allowed correlation of carbonyl (C-13) carbons with directly bonded C-13 alpha and N-15 spins in ARD (Kostic, M.; Pochapsky, S. S.; Pochapsky, T. C. J. Am. Chem. Soc. 2002, 124,9054-9055), but not N-15 with C-13 alpha, a critical connection for sequential assignment of backbone resonances. It is now shown that complete sample deuteration combined with direct C-13 detection using a cold probe/preamplifier permits the one-bond C-13 alpha-N-15 correlation to be made via a four-pulse double-quantum experiment CAN. Combined with data from other C-13 direct-observe 20 NMR experiments, CAN data permits sequential assignments to be made for many resonances in ARD close to the active-site metal.