Properties of 2'5' oligoadenylate synthetase.
2'5' Oligoadenylate synthetase has been purified from rabbit reticulocyte lysate to a specific activity of 37 units per mg protein (1 unit incorporates 1 nmole of AMP residue into oligoadenylate per min at 37 degrees C) by DEAE-cellulose and rI:rC-agarose chromatography and (NH4)2SO4 precipitations. For this a rapid and quantitative assay was developed, based on TLC with PEI-cellulose. In a single step ATP and the 2'5' oligoadenylates (pppA(2p5A)n 1 less than n less than 14) are separated. The enzyme showed the following properties: a pH optimum around 8, a Mg++ requirement with full activation at 20 mM, no effect of KCl between 25 and 100 mM but complete inactivation at 120 mM, 30% stimulation by 10% ethanol, and a half-life at 52 degrees C of around 5 min. The maximum yield of the reaction ws 95% conversion of ATP into 2'5' oligoadenylates. The mechanism of elongation is not processive. Kinetic studies on the formation of different oligomeric intermediates suggest a mechanism of a dissipative nature. The products are mainly dimers, trimers and tetramers. The longest oligomer seen had about 15 AMP residues. The enzyme has no absolute substrate specificity for ATP, GTP, or UTP can also be incorporated, in the presence of ATP, by the enzyme into a co-oligonucleotide containing A and G or A and U. The enzyme can also add one unit of GMP or UMP onto a primer 2'5'pppApA or pppApApA.
D015088:2',5'-Oligoadenylate Synthetase; D000227:Adenine Nucleotides / Q000096:biosynthesis; D000255:Adenosine Triphosphate; D000818:Animals; D006160:Guanosine Triphosphate; D066298:In Vitro Techniques; D007700:Kinetics; D009843:Oligoribonucleotides / Q000096:biosynthesis; D011118:Polynucleotide Ligases / Q000302:isolation & purification / Q000378:metabolism*; D011817:Rabbits; D013379:Substrate Specificity; D014544:Uridine Triphosphate
D000227:Adenine Nucleotides; D009843:Oligoribonucleotides; C023505:2',5'-oligoadenylate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D015088:2',5'-Oligoadenylate Synthetase; D011118:Polynucleotide Ligases; D014544:Uridine Triphosphate