Applicability of the phosphorus-31 (oxygen-17) nuclear magnetic resonance method in the study of enzyme mechanism involving phosphorus

Abstract

The [31P (17O)] NMR method is tested for its general applicability by correlation with the line widths of the 17O NMR signals for the following compounds: trimethyl [17O4]phosphate (1), [17O4]phosphate (2), [.alpha.-17O2]adenosine 5''-(thiophosphate) (3), [.alpha.-17O,.alpha..beta.-17O]adenosine 5''-(1-thiotriphosphate) (4), [.gamma.-17O3]ATP (5), [.alpha..beta.,.beta..gamma.-17O2,.beta.-17O2]ATP (6), Mg [.gamma.-17O3]ATP (7), and Mg [.alpha..beta.,.beta..gamma.-17O2,.beta.-17O2]ATP (8). The line broadening effect of 17O on the 31P signals of the directly bonded 31P nuclei is present for all the functional groups in the above compounds which include examples of small 17O line widths (1 and 2), of intermediate 17O line widths (3, 5, and the nonbridge 17O of 4 and 6), and of very large 17O line widths (7, 8 and the bridge 17O of 4 and 6). On the basis of the established approximate relationship .DELTA.X.DELTA.Q .apprxeq. aJ2, where .DELTA.X and .DELTA.Q are the 31P and 17O line widths, respectively, of the 31P-17O groups, a is a constant, and J is the 31P-17O coupling constant, the results suggest that for most of the phosphate derivatives of biochemical interest, the line broadening effect of 17O should be present and detectable; i.e., .DELTA.X should be larger than the limiting value (.apprx. 20 HZ). Mg2+ causes the 17O signal of 5 and 6 to broaden (.DELTA.Q increases), which in turn causes the 31P signal to sharpen (.DELTA.X decreases). The 31P (17O) NMR method, in combination with 17O NMR, could become a tool to study diamagnetic metal ion-nucleotide interactions.