Robert C. Hodson

Preparation of adenosine-3′-phosphate-5′-phosphosulfate (PAPS): An improved enzymatic method using Chlorella pyrenoidosa

Abstract Adenosine-3′-phosphate-5′-phosphosulfate (PAPS) is prepared from crude French press extracts of the green alga Chlorella pyrenoidosa purified somewhat by high-speed centrifugation and brief dialysis. The PAPS synthesized in an overnight incubation from sulfate and ATP is purified by Dowex-1 (Cl − ) ion-exchange chromatography and adsorption on activated charcoal. Incubations with carrier-free sulfate routinely yield PAPS with 60–80% of the initial radioactivity and a radiochemical purity exceeding 90%. Any desired specific activity may be obtained by diluting the radiosulfate with nonradioactive sulfate, and the method may be used equally well for the preparation of nonradioactive PAPS. For routine use we prepare PAPS from sulfate- 35 S having a specific activity of about 1–2 mCi/mmole. A radiochemical yield of 10–15% is obtained, representing 180–270 μmoles of PAPS. The nucleotide and radiochemical purities of this material are 90% or better, and sulfate- 35 S is the only detectable radioactive contaminant.