Phosphorus-31 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(II) 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 degrees C to determine the effect of exchange on the observed relaxation rates. Relaxation rates in the E.MnATP, E.MnGTP, and E.MnGDP complexes were shown to be exchange-limited and therefore without structural information. Relaxation rates for the complexes E.CoATP, E.CoGTP, and E.CoGDP were shown to depend on Co(II)-31P distances. Inability to precisely estimate spectral densities arising from electronic relaxation of Co(II) restricts calculations of Co(II)-31P distances in these complexes to upper and lower limits. At the center of these limits, the Co(II)-31P distances of beta-P and gamma-P in E.CoATP and E.CoGTP, and of beta-P (E.CoGDP), are in the range 3.1-3.5 A appropriate for the first coordination sphere. For all these complexes, the corresponding distance for alpha-P is appreciably larger in the range 3.9-4.5 A.(ABSTRACT TRUNCATED AT 250 WORDS)

Associative mechanism for phosphoryl transfer: A molecular dynamics simulation of Escherichia coli adenylate kinase complexed with its substrates

“…In porcine muscle AK · CoATP, AK · CoGTP, and AK · CoGDP, 31 P NMR relaxation rates indicate that Co 2+ is directly coordinated to only β‐PO 4 (GDP) or β‐PO 4 and γ‐PO 4 (ATP and GTP) 54. In addition, α‐PO 4 is too far from Co 2+ to be in the first coordination sphere, but also too close to be in the second coordination sphere (not enough room for a water molecule).…”

Investigations of kinase substrate specificity with aqua rhodium(III) complexes of adenosine 5'-triphosphate

“…The structure of MgATP bound in the active site of myokinase has also been examined using several techniques other than the exchange-inert metal-ATP technique. Specifically, on the basis of 31P spin-relaxation measurements, Ray et al (1988) concluded that the structure of MgATP bound in the myokinase active site is /3,7-bidentate, and the results obtained from Mg(II)/Cd(II) ATP/3S/ATPaS studies (Tomasselli & Noda, 1981Tomasselli et al, 1984) and Mn-170-ATP EPR studies (Kalbitzer et al, 1983) indicate that the configuration at the /3-P is . Thus, the results from each of these studies provide evidence for /3,7-bidentate Mg(H20)4ATP as the myokinase substrate.…”

Conformation of Manganese(II)−Nucleotide Complexes Bound to Rabbit Muscle Creatine Kinase: 13C NMR Measurements Using [2-13C]ATP and [2-13C]ADP

“…In the case of adenylate kinase, however, the data indicate β,γbidentate binding of the cation in the enzyme-metal-ATP complex, and β-monodentate binding in the enzyme-metal-GDP complex. The distance of R-P to the cation is ∼1.0 Å longer than that for direct coordination in both cases(Ray et al, 1988).7240 Biochemistry, Vol. 35, No.…”

“…The theory of nuclear spin relaxation in the presence of paramagnetic cations is well-known (Dwek, 1973;James, 1973;Mildvan & Gupta 1978;Burton et al, 1979;Jardetzky & Roberts, 1981). A summary of this theory and the experimental strategy arising from it have previously been published (Jarori et al, 1985(Jarori et al, , 1989Ray & Nageswara Rao, 1988b;Ray et al, 1988). The relevant equations for the analysis of the data in this paper are given below.…”

“…31 P spinrelaxation measurements in enzyme-bound nucleotide complexes of several kinases in the presence of Co(II) have been used to establish that the cation is directly coordinated to the phosphate groups. The enzymes studied include creatine kinase (Jarori et al, 1985), 3-phosphoglycerate kinase (Ray & Nageswara Rao, 1988a,b), adenylate kinase 2 (Ray et al, 1988), and arginine kinase (Jarori et al, 1989). Proton TRNOESY measurements have been used to deduce the adenosine conformation of MgATP and MgADP bound to creatine kinase (Murali et al, 1993), arginine kinase (Murali et al, 1994), and pyruvate kinase (Jarori et al, 1994) and of MgATP bound to the adenyl transfer enzyme, methionyl tRNA synthetase (N. Murali, Y. Lin, Y. Mechulam, P. Plateau, and B. D. Nageswara Rao, submitted), and to the pyrophosphoryl transfer enzyme phosphoribosylpyrophosphate (PRPP) synthetase (Jarori et al, 1995).…”