Manganese complexes nucleotides

Figure 4 Ribbon diagrams of glycosyltransferases that demonstrate the different structural folds. When possible, nucleotide-donor sugars are depicted as stick models, manganese is depicted by a magenta ball, and N- and C-terminals are labeled. GT-A fold members are represented by bovine pi,4-galactosyltransferase I complexed with UDP-Gal (5) (Fig. (4)a, PDB accession number 1O0R) and by human polypeptide...

Kinetic studies of NMP kinases, as well as many other enzymes having ATP or other nucleoside triphosphates as a substrate, reveal that these enzymes are essentially inactive in the absence of divalent metal ions such as magnesium (Mg2+) or manganese (Mn2+), but acquire activity on the addition of these ions. In contrast with the enzymes discussed so far, the metal is not a component of the active site. Rather, nucleotides such as ATP bind these ions, and it is the metal ion-nucleotide complex that is the true substrate for the enzymes. The dissociation constant for the ATP-Mg2+ complex is approximately 0.1 mM, and thus, given that intracellular Mg + concentrations are typically in the millimolar range, essentially all nucleoside triphosphates are present as NTP-Mg + complexes. 

The chemical shift of the phosphorus resonance of various nucleotides has been studied as a function of pH in the presence and absence of RNase A. The signal shifts upheld on protonation of the phosphate and the apparent pATa of the phosphate group in 2 -CMP complex with RNase is the same as the pATa of histidine-119 in this enzyme as determined by n.m.r. From n.m.r. relaxation rates for the ternary complex manganese(n)-phosphate-E. coli alkaline phosphatase, it has been concluded that an outer-sphere complex is formed which has a shorter lifetime than the enzyme turnover rate. The latter conclusion is consistent with the participation of the complex in the enzymic reaction. 

The coordination structures of enzyme-bound manganese nucleotides can in favorable cases be determined by analysis of electron paramagnetic resonance (EPR) spectra of Mn(II) coordinated to O-labeled nucleotides. When the nucleotide is stereospecifically labeled with O at one diastereotopic position of a prochiral center, either oxygen can in principle be bound to Mn(II) in the coordination complex in an enzymic site. When the coordination bond is between Mn(II) and O, the EPR signals for Mn(II) are broadened and attenuated, owing to unresolved superhyperfine coupling between the nucleus of 0 and the unpaired electrons of Mn(II) (23). No such effect is possible with 0, which has no nuclear spin. The effect is observable in samples in which all the Mn(Il) is specifically bound in one or two defined complexes of the nucleotide with the enzyme. Thus the complex Mg(Sp)-[a- 0]ADP bound at the active site of ere-... 

The coordination structures of manganese nucleotides at enzymic active sites are especially good models for magnesium nucleotides, since they are comparably reactive as substrates. The coordination exchange-inert complexes are somewhat less reactive, but no conflicting results have been obtained to date by these three methods. 

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