Purification, properties and substrate specificity of adenosine triphosphate sulphurylase from spinach leaf tissue.

摘要: 1. ATP sulphurylase was purified up to 1000-fold from spinach leaf tissue. Activity measured by sulphate-dependent [32P]PPi–ATP exchange. The enzyme separated Mg2+-requiring alkaline pyrophosphatase (which interferes with the PPi–ATP-exchange assay) and other activities. No ADP activity detected. 2. Sulphate only form of inorganic sulphur that catalysed PPi–ATP exchange; Km (sulphate) 3.1mm, (ATP) 0.35mm pH optimum 7.5–9.0. insensitive thiol-group reagents required either Mg2+ or Co2+ for activity. 3. [32P]PPi–dATP (dATP) 0.84mm V 30% (ATP). Competition between dATP demonstrated. 4. Selenate exchange competed sulphate; (selenate) 1.0mm (sulphate). AMP formed selenate as substrate. Molybdate did not catalyse exchange, but formed. 5. Synthesis adenosine 5′-[35S]sulphatophosphate demonstrated coupling Mg2+-dependent (also prepared spinach) [35S]sulphate substrates; 5′-sulphatophosphate synthesized in absence pyrophosphatase. Some parameters coupled system are reported.