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1. An F−-insensitive 3′-nucleotidase was purified from spinach leaf tissue; the enzyme hydrolysed 3′-AMP, 3′-CMP and adenosine 3′-phosphate 5′-sulphatophosphate but not adenosine 5′-nucleotides nor PPi. The pH optimum of the enzyme was 7.5; Km (3′-AMP) was approx. 0.8mm and Km (3′-CMP) was approx. 3.3mm. 3′-Nucleotidase activity was not associated with chloroplasts. Purified Mg2+-dependent pyrophosphatase, free from F−-insensitive 3′-nucleotidase, catalysed some hydrolysis of 3′-AMP; this activity was F−-sensitive. 2. Adenosine 5′-sulphatophosphate kinase activity was demonstrated in crude spinach extracts supplied with 3′-AMP by the synthesis of the sulphate ester of 2-naphthol in the presence of purified phenol sulphotransferase; purified ATP sulphurylase and pyrophosphatase were also added to synthesize adenosine 5′-sulphatophosphate. Adenosine 5′-sulphatophosphate kinase activity was associated with chloroplasts and was released by sonication. 3. Isolated chloroplasts synthesized adenosine 3′-phosphate 5′-sulphatophosphate from sulphate and ATP in the presence of a 3′-nucleotide; the formation of adenosine 5′-sulphatophosphate was negligible. In the absence of a 3′-nucleotide the synthesis of adenosine 3′-phosphate 5′-sulphatophosphate was negligible, but the formation of adenosine 5′-sulphatophosphate was readily detected. Some properties of the synthesis of adenosine 3′-phosphate 5′-sulphatophosphate by isolated chloroplasts are described. 4. Adenosine 3′-phosphate 5′-sulphatophosphate, synthesized by isolated chloroplasts, was characterized by specific enzyme methods, electrophoresis and i.r. spectrophotometry. 5. Isolated chloroplasts catalysed the incorporation of sulphur from sulphate into cystine/cysteine; the incorporation was enhanced by 3′-AMP and l-serine. It was concluded that adenosine 3′-phosphate 5′-sulphatophosphate is an intermediate in the incorporation of sulphur from sulphate into cystine/cysteine.

1. Extracts of human peripheral blood lymphocytes contained a histone phosphatase that catalysed the release of P(i) from phosphorylated whole thymus histone. 2. Stimulation of the phosphatase was obtained by concentrations of KCl and NaCl of up to 75mm, and by MgCl(2); CaCl(2) inhibited the enzymic activity. 3. In the absence of MgCl(2), phosphoenol-pyruvate inhibited histone phosphatase activity; this inhibition could be partially reversed by adding MgCl(2) to assays. 4. Lymphocyte extracts contained a protein kinase activity which was maximally stimulated by 1mum-cyclic AMP (adenosine 3':5'-cyclic monophosphate) or by 0.1mm-cyclic GMP (guanosine 3':5'-cyclic monophosphate). 5. Incubation of the enzyme with histone in the absence of ATP or MgCl(2) resulted in the dissociation of the enzyme into a lower-molecular-weight species that was not stimulated by cyclic AMP. This effect could be prevented if ATP and MgCl(2) were present in reaction mixtures before histone and enzyme were allowed to interact. 6. Cyclic AMP also dissociated the kinase into a lower-molecular-weight species. 7. In the presence of 1mum-AMP, half-maximal activities were obtained with 0.92mm-MgCl(2), 6.0mum-ATP and 0.23mg of whole thymus histone/ml.

Nuclei isolated from glucagon-stimulated perfused rat livers contained 2–3 times as much protein kinase activity as did nuclei from control animals. In the presence of either the heat-stable inhibitor or the protein kinase regulatory subunit the elevated cyclic AMP-independent enzyme activity from stimulated nuclei was inhibited to an activity equivalent to that found in controls.