31P NMR Studies of the Structure of Cation-Nucleotide Complexes Bound to Porcine Muscle Adenylate Kinase?

ABSTRACT: The paramagnetic effects on the spin-relaxation rates of 31P nuclei in complexes of porcine muscle adenylate kinase with ATP, GTP, GDP, and AMP were measured in the presence of two dissimilar activating paramagnetic cations, Mn(I1) and Co(II), to examine the structures of the enzyme-bound complexes. Experiments were performed exclusively on enzyme-bound complexes to limit contributions to observed relaxation rates to two exchanging complexes (with and without cation). Measurements were made at three frequencies, 81, 121.5, and 190.2 MHz, and as a function of temperature in the range 5-30 OC to determine the effect of exchange on the observed relaxation rates. Relaxation rates in the E-MnATP, E-MnGTP, and ESMnGDP complexes were shown to be exchange-limited and therefore without structural information. Relaxation rates for the complexes ECoATP, ECoGTP, and E-CoGDP were shown to depend on C O ( I I)- ~ ~ P distances. Inability to precisely estimate spectral densities arising from electronic relaxation of Co(I1) restrict