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. 1973 Jul;133(3):541–550. doi: 10.1042/bj1330541

Adenosine 5′-triphosphate sulphurylase from Saccharomyces cerevisiae

Catherine S Hawes 1, D J D Nicholas 1
PMCID: PMC1177733  PMID: 4582048

Abstract

1. ATP sulphurylase from Saccharomyces cerevisiae was purified 140-fold by using heat treatment, DEAE-cellulose chromatography and Sepharose 6B gel filtration. 2. The enzyme was stable at −15°C, optimum reaction velocity was between pH7.0 and 9.0, and the activation energy was 62kJ/mol (14.7kcal/mol). 3. The substrate was shown to be the MgATP2− complex, free ATP being inhibitory. 4. Double-reciprocal plots from initial-velocity studies were intersecting and the Km of each substrate was determined at infinite concentration of the other (Km MgATP2−, 0.07mm; MoO42−, 0.17mm). 5. Radio-isotopic exchange between the substrate pairs, adenosine 5′-[35S]sulphatophosphate and SO42−, 35SO42− and adenosine 5′-sulphatophosphate, occurred only in the presence of either MgATP2− or PPi. This suggests, along with the initial-velocity data, a sequential reaction mechanism in which both substrates bind before any product is released. 6. The enzyme reaction was specific for ATP and was not inhibited by l-cysteine, l-methionine, SO32−, S2O32− (all 2mm) nor by p-chloromercuribenzoate (1mm). 7. Competitive inhibition of the enzyme with respect to MoO42− was produced by SO42− (Ki=2.0mm) and non-competitive inhibition by sulphide (Ki=3.4mm). 8. Adenosine 5′-sulphatophosphate inhibited strongly and concentrations as low as 0.02mm altered the normal hyperbolic velocity–substrate curves with both MgATP2− and MoO42− to sigmoidal forms.

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Selected References

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