MULTIPLE IONIZATIONS IN PROTEINS - ARSENICAL ANALOGS OF NATURAL PHOSPHATES

1. Handling multiple ionizations is facilitated by realizing that the titration of a substance with n ionizing groups is normally the sum of n one-site curves; the dissociation constants characterizing these cu rves are 'titration' constants. The pH dependence of any property is t he sum of one-site curves with these constants. Each group in the mole cule titrates in fractions, with a fraction titrating with each titrat ion constant. 2. When enzymes that esterify, acylate or phosphorylate the -O-PO3H2 group of substrates act on analogues in which this group is replaced with -CH2-AsO3H2, spontaneous hydrolysis follows the enzym e-catalysed reaction. This is because water easily replaces the alkoxy and acyloxy groups on arsenic that such enzymes introduce. Thus RNA p olymerase and adenylate kinase effectively exhibit exonuclease and ATP ase activities when given these analogues of diphosphate and AMP, resp ectively. A new consequence of enzymic transformations of such analogu es is observed when glycerol-phosphate dehydrogenase oxidizes HO-CH2-C H(OH)-CH2-CH2-AsO3H2, and when alcohol dehydrogenase oxidizes HO-CH2-C H2-CH2-AsO3H2; the oxidation is followed by arsenite release, presumab ly a consequence of enolization of the ketone first formed.