New assay for ATP sulfurylase based on differential solubility of the sodium salts of adenosine 5'-phosphosulfate and sulfate. [/sup 35/S tracer techniques]
A new assay for ATP sulfurylase (ATP:sulfate adenyltransferase, EC 2.7.7.4) has been devised, the key feature of which is the quantitative precipitation of the unreacted substrate as Na/sub 2/SO/sub 4/, in ethanol:water (5:1), leaving the product adenosine 5'-phosphosulfate (APS) in solution with an 80 percent yield. This separation procedure, coupled with the use of (/sup 35/S)SO/sub 4//sup 2 -/, makes it possible to assay the synthesis of picomole amounts of (/sup 35/S)APS and thereby determine accurately the initial velocity of the forward reaction catalyzed by ATP sulfurylase. Extracts from tobacco cells synthesized (/sup 35/S)APS and a negligible amount of one unknown sulfur compound, but they did not synthesize 3'-phosphoadenosine 5'-(/sup 35/S)phosphosulfate (PAPS) under the assay conditions adopted, so that the /sup 35/S remaining in solution after ethanol precipitation of (/sup 35/S)SO/sub 4//sup 2 -/ is a valid assay of the ATP sulfurylase reaction in these extracts. This assay is simple, rapid, and suitable for assaying multiple samples. Extracts of four additional species of plants were incubated under the same conditions, and each catalyzed the synthesis of (/sup 35/S)APS according to this assay. The assay is suitable for use with the enzyme from any organism, provided that the product APS is not acted upon by another enzyme.
- Research Organization:
- Michigan State Univ., East Lansing
- OSTI ID:
- 7314611
- Journal Information:
- Anal. Biochem.; (United States), Journal Name: Anal. Biochem.; (United States) Vol. 75:2; ISSN ANBCA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ADENOSINE
ALKALI METAL COMPOUNDS
ATP
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOASSAY
DAYS LIVING RADIOISOTOPES
ENZYMES
EVEN-ODD NUCLEI
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
NUCLEI
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
OXYGEN COMPOUNDS
PHOSPHORUS COMPOUNDS
PLANT CELLS
RADIOISOTOPES
RIBOSIDES
SODIUM COMPOUNDS
SOLUBILITY
SULFATES
SULFUR 35
SULFUR COMPOUNDS
SULFUR ISOTOPES
TOBACCO
TRACER TECHNIQUES
TRANSFERASES