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[Substrate specificity of T4 RNA-ligase: the role of a purine nucleotide base in forming a covalent AMP-RNA-ligase

B A Iuodka, D Ia Labeĭkite, S I Sasnauskene

Biokhimiia (Moscow, Russia)

|June 1, 1993

Abstract:

The NTP binding site of bacteriophage T4 RNA-ligase (EC 6.5.1.3) was studied using several ATP analogs modified in the purine moiety of the nucleotide at positions 1, 2, 6 and 8: adenosine-N'-oxide 5'-triphosphate (I), 6-chloropurine riboside 5'-triphosphate (II), 6-mercaptopurine riboside 5'-triphosphate (III), 1,N6-ethenoadenosine 5'-triphosphate (IV), 8-sulphoadenosine 5'-triphosphate (V), 8-bromoadenosine 5'-triphosphate (VI), inosine 5'-triphosphate (VII) and guanosine 5'-triphosphate (VIII). For all of the ATP analogs under study a reversible inhibition was demonstrated. These analogs appeared to be competitive inhibitors of the T4 RNA-ligase adenylation reaction and only V, VI and VII were efficient as substrates. The kinetic parameters for the competitive inhibition, (I50, Ki) were determined. A putative structure for the T4 RNA-ligase active site was proposed.

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