Labeled adenosine(5′)tetraphospho(5′)adenosine (Ap4A) and adenosine(5′)tetraphospho(5′)nucleoside (Ap4N). Synthesis with firefly luciferase

Garrido, Sofía; Zaera, E.; A, Torrecilla; Sillero, Antonio; Sillero, Marı́a A. Günther

doi:10.1016/0165-022x(95)00007-e

Cited by 7 publications

(2 citation statements)

References 10 publications

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“…The dipeptides were detected in solution as ‘shunt’ products and not as enzyme bound intermediates. Some members of the adenylate forming superfamily, such as the acyl CoA synthetases [22] and luciferase [23,24], catalyze in the presence of pyrophosphatase and ATP or polyphosphates the formation of a variety of adenosine 5′‐polyphosphates (Ap n A and p n A) by transferring the adenylyl moiety from the aminoacyl adenylate onto the corresponding acceptor. The results support the occurrence of two binding sites for ATP, one with high affinity for MgATP 2− and high specificity to form an E·AA~AMP complex and the other with a low affinity and low specificity, binding ATP, NTP and polyphosphate chains.…”

Section: Discussionmentioning

confidence: 99%

Active site titration of gramicidin S synthetase 2: evidence for misactivation and editing in non‐ribosomal peptide biosynthesis

Kittelberger¹,

Pavela-Vrančić²,

Döhren³

1999

FEBS Letters

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The catalytic competence of gramicidin S synthetase 2 (GS2) was determined by following the kinetics of PP i generation using active site titration measurements with [Q Q-32 P]ATP. The initial`burst' of product formation can be correlated to the generation of the aminoacyl adenylate:enzyme complexes at the four amino acid activation domains and the subsequent aminoacylation of carrier domains, followed by a slow linear turnover of substrate due to breakdown of the intermediate. Simultaneous activation of all four amino acid substrates at a saturating concentration displayed a consumption of 8.3 ATP/GS2. In the presence of single amino acids, a binding stoichiometry higher than the anticipated two ATP per active site was obtained, implying misactivation at non-cognate domains. Breakdown of acyladenylate intermediates reflects a possible corrective mechanism by which the enzyme controls the fidelity of product formation.z 1999 Federation of European Biochemical Societies.

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Section: Discussionmentioning

confidence: 99%

Active site titration of gramicidin S synthetase 2: evidence for misactivation and editing in non‐ribosomal peptide biosynthesis

Kittelberger¹,

Pavela-Vrančić²,

Döhren³

1999

FEBS Letters

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“…[18]. However when the ATP concentration in the assay is very low (M), the enzyme behaves as an ATPase liberating AMP[29,30]. The synthesis of ATP mevalonate derivatives was approached here, incubating luciferase in the presence of 0.02 mM [-32 P]ATP and iso-PPP, far-PPP or ger-PPP at 0.3 mM each (Fig.…”

mentioning

confidence: 99%

Synthesis of ATP derivatives of compounds of the mevalonate pathway (isopentenyl di- and triphosphate; geranyl di- and triphosphate, farnesyl di- and triphosphate, and dimethylallyl diphosphate) catalyzed by T4 RNA ligase, T4 DNA ligase and other ligases

Sillero¹,

Diego²,

Tavares³

et al. 2009

Biochemical Pharmacology

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Compounds of the mevalonate pathway containing a terminal di- or triphosphate (mev-PP or mev-PPP) were tested as substrates of several enzyme ligases (T4 RNA ligase, T4 DNA ligase, firefly luciferase and other ligases) for the synthesis of ATP derivatives of the mev-pppA or mev-ppppA type. T4 RNA ligase, in the presence of ATP and the substrates: geranyl, farnesyl or isopentenyl triphosphates, and geranyl, farnesyl, dimethylallyl or isopentenyl diphosphates, all at 0.3 mM concentration, catalyzed the synthesis of the corresponding ATP derivatives at a relative rate of activity of: 7.6+/-1.4 mU/mg or 100%; 39%; 42%; 24%; 18%; 12% and 6%, respectively. Inhibition (%) of the synthesis by excess of substrate (0.8 mM vs. 0.3 mM) was observed with farnesyl diphosphate (99%); farnesyl triphosphate (96%) and geranyl triphosphate (32%). V(max), K(m), K(cat) and K(cat)/K(m) values were also determined. The K(cat)/K(m) values calculated were for: farnesyl triphosphate, 166; geranyl triphosphate, 52.2; farnesyl diphosphate, 12.1; geranyl diphosphate, 8.6; isopentenyl triphosphate, 6.7; dimethylallyl diphosphate, 3.1 and isopentenyl diphosphate, 0.9. Similar results were obtained with T4 DNA ligase. The above-mentioned compounds were also substrates of firefly luciferase synthesizing the mev-pppA or mev-ppppA derivatives. In our hands, neither the acyl- or acetyl-CoA synthetases nor the ubiquiting activating enzyme (E1) catalyzed the synthesis of ATP derivatives of these compounds. The results here presented could be related with the mechanism of action of bisphosphonates on osteoclasts or tumor cells.

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Selective Degradation of 2′-Adenylated Diadenosine Tri- and Tetraphosphates, Ap3A and Ap4A, by Two Specific Human Dinucleoside Polyphosphate Hydrolases

Guranowski

Galbas

Hartmann

et al. 2000

Archives of Biochemistry and Biophysics

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Labeled adenosine(5′)tetraphospho(5′)adenosine (Ap4A) and adenosine(5′)tetraphospho(5′)nucleoside (Ap4N). Synthesis with firefly luciferase

Cited by 7 publications

References 10 publications

Active site titration of gramicidin S synthetase 2: evidence for misactivation and editing in non‐ribosomal peptide biosynthesis

Active site titration of gramicidin S synthetase 2: evidence for misactivation and editing in non‐ribosomal peptide biosynthesis

Synthesis of ATP derivatives of compounds of the mevalonate pathway (isopentenyl di- and triphosphate; geranyl di- and triphosphate, farnesyl di- and triphosphate, and dimethylallyl diphosphate) catalyzed by T4 RNA ligase, T4 DNA ligase and other ligases

Selective Degradation of 2′-Adenylated Diadenosine Tri- and Tetraphosphates, Ap3A and Ap4A, by Two Specific Human Dinucleoside Polyphosphate Hydrolases

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