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Adenylylsulfate-ammonia adenylyltransferase activity is another inherent property of Fhit proteins.

ABSTRACT: Fhits (fragile histidine triad proteins) occur in eukaryotes but their function is largely unknown, although human Fhit is believed to act as a tumour suppressor. Fhits also exhibit dinucleoside triphosphatase, adenylylsulfatase and nucleoside phosphoramidase activities that in each case yield nucleoside 5'-monophosphate as a product. Due to the dinucleoside triphosphatase activity, Fhits may also be involved in mRNA decapping. In the present study, we demonstrate Fhit-catalysed ammonolysis of adenosine 5'-phosphosulfate, which results in the formation of adenosine 5'-phosphoramidate. This reaction has previously been associated with adenylylsulfate-ammonia adenylyltransferase (EC 2.7.7.51). Our finding shows that the capacity to catalyse ammonolysis is another inherent property of Fhits. Basic kinetic parameters and substrate specificity of this reaction catalysed by human Fhit are presented.

SUBMITTER: Wojdyla-Mamon AM

PROVIDER: S-EPMC4613722 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Adenylylsulfate-ammonia adenylyltransferase activity is another inherent property of Fhit proteins.

Wojdyła-Mamoń Anna M AM Guranowski Andrzej A

Bioscience reports 20150625 4

Fhits (fragile histidine triad proteins) occur in eukaryotes but their function is largely unknown, although human Fhit is believed to act as a tumour suppressor. Fhits also exhibit dinucleoside triphosphatase, adenylylsulfatase and nucleoside phosphoramidase activities that in each case yield nucleoside 5'-monophosphate as a product. Due to the dinucleoside triphosphatase activity, Fhits may also be involved in mRNA decapping. In the present study, we demonstrate Fhit-catalysed ammonolysis of a ...[more]

PMID: 26181368

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Project description:Extracts of Chlorella pyrenoidosa, Euglena gracilis var. bacillaris, spinach, barley, Dictyostelium discoideum and Escherichia coli form an unknown compound enzymically from adenosine 5'-phosphosulphate in the presence of ammonia. This unknown compound shares the following properties with adenosine 5'-phosphoramidate: molar proportions of constituent parts (1 adenine:1 ribose:1 phosphate:1 ammonia released at low pH), co-electrophoresis in all buffers tested including borate, formation of AMP at low pH through release of ammonia, mass and i.r. spectra and conversion into 5'-AMP by phosphodiesterase. This unknown compound therefore appears to be identical with adenosine 5'-phosphoramidate. The enzyme that catalyses the formation of adenosine 5'-phosphoramidate from ammonia and adenosine 5'-phosphosulphate was purified 1800-fold (to homogeneity) from Chlorella by using (NH(4))(2)SO(4) precipitation and DEAE-cellulose, Sephadex and Reactive Blue 2-agarose chromatography. The purified enzyme shows one band of protein, coincident with activity, at a position corresponding to 60000-65000 molecular weight, on polyacrylamide-gel electrophoresis, and yields three subunits on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of 26000, 21000 and 17000 molecular weight, consistent with a molecular weight of 64000 for the native enzyme. Isoelectrofocusing yields one band of pI4.2. The pH optimum of the enzyme-catalysed reaction is 8.8. ATP, ADP or adenosine 3'-phosphate 5'-phosphosulphate will not replace adenosine 5'-phosphosulphate, and the apparent K(m) for the last-mentioned compound is 0.82mm. The apparent K(m) for ammonia (assuming NH(3) to be the active species) is about 10mm. A large variety of primary, secondary and tertiary amines or amides will not replace ammonia. One mol.prop. of adenosine 5'-phosphosulphate reacts with 1 mol.prop. of ammonia to yield 1 mol.prop. each of adenosine 5'-phosphoramidate and sulphate; no AMP is found. The highly purified enzyme does not catalyse any of the known reactions of adenosine 5'-phosphosulphate, including those catalysed by ATP sulphurylase, adenosine 5'-phosphosulphate kinase, adenosine 5'-phosphosulphate sulphotransferase or ADP sulphurylase. Adenosine 5'-phosphoramidate is found in old samples of the ammonium salt of adenosine 5'-phosphosulphate and can be formed non-enzymically if adenosine 5'-phosphosulphate and ammonia are boiled. In the non-enzymic reaction both adenosine 5'-phosphoramidate and AMP are formed. Thus the enzyme forms adenosine 5'-phosphoramidate by selectively speeding up an already favoured reaction.

Dataset Information

Adenylylsulfate-ammonia adenylyltransferase activity is another inherent property of Fhit proteins.

Publications

Adenylylsulfate-ammonia adenylyltransferase activity is another inherent property of Fhit proteins.

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