Dextran-linked purine nucleosides as substrates and inhibitors of adenosine deaminase.

Dextran-bound adenosine, inosine, and nebularine have been prepared by carbodiimide coupling of their 2',3'-O-(4-carboxyethyl-1-methylbutylidene) cyclic acetal derivatives to 6-aminohexyldextran or 12-aminododecanyldextran. The latter polymers were prepared by cyanogen-bromide activation of dextran T80 followed by reaction with 1,6-diaminohexane or 1,12-diaminododecane. A high CNBr concentration leads to high-molecular-weight material, probably due to cross-linking, accompanied by a decrease in the digestion velocity using endo-dextranase from Penicillium species (EC 3.2.1.11). The dextran-bound nucleosides, as well as the nucleoside 2',3'-O-(4-ethoxycarbonyl-1-methylbutylidene) acetal derivatives, were tested as substrates and inhibitors for adenosine deaminase. The Km of the adenosine acetal ester is identical to that of adenosine which shows that acetalation does not hinder complex formation. Since the maximum velocity of deamination is decreased fourfold, the modified substrate does not fit as well as the nucleoside. The polymer-bound acetals show a 3-8-fold increase of Km or Ki and unchanged V compared to the corresponding acetals while dextranase digestion of the support does not alter the kinetic data. This indicates that the length of the polysaccharide chain does not interfere either with the complex formation or with the catalytic activity of the modified substrate. Since the activation energies of the deamination reactions of adenosine, its acetal ester, and dextran-linked adenosine are all similar (29.8-32.3 kJ mol-1) it is concluded that no diffusion control of the enzymatic reaction results from the binding of the nucleoside acetals to dextran T80.