Metal‐directed affinity labelling

Dahl, Knut Helkås; McKinley-McKee, John S.; Beyerman, H. C.; Noordam, A.

doi:10.1016/0014-5793(79)80979-5

Cited by 13 publications

(5 citation statements)

References 12 publications

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“…This phosphatase binds Zn2+ with a /if of 108-10m M~' (Friedberg, 1974). The ATP splitting activity is not inhibited by 4 mM of the so-called metal-directed affinity labeling reagent (7?,S)-2-bromo-3- (5-imidazoyl)propionic acid (Dahl et al, 1979).…”

Section: Resultsmentioning

confidence: 99%

A novel enzymic activity of phenylalanyl transfer ribonucleic acid synthetase from baker's yeast: zinc ion induced transfer ribonucleic acid independent hydrolysis of adenosine triphosphate

Igloi¹,

Haar²,

Cramer³

1980

Biochemistry

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Phenylalanyl-tRNA synthetase from baker's yeast in the presence of phenylalanine or other amino acids misactivated by the enzyme, ATP, and low concentrations of Zn2+ is able to hydrolyze ATP to AMP and PPi very efficiently. After dialysis of the enzyme against ethylenediaminetetraacetic acid (EDTA), this amino acid dependent but tRNAPhe-independent hydrolysis is suppressed to negligible levels. The ATP hydrolysis can be restored by the addition of Zn2+ to the EDTA-dialyzed enzyme. During aminoacylation of tRNAPhe the Zn2+-induced ATP hydrolysis parallels the aminoacylation reaction, leading to nonstoichiometric production of AMP. Mechanistically, we conclude that Zn2+ can be bound to phenylalanyl-tRNA synthetase and can influence the stability of ATP if an activatable amino acid is present. The influence of Zn2+, if any, on the aminoacylation of tRNAPhe is not known. In practice, this side reaction is of the utmost importance in all cases in which the fate of ATP during aminoacylation is followed, especially if the stoichiometry of ATP consumption in relation to Phe-tRNAPhe formation has to be determined.

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Section: Resultsmentioning

confidence: 99%

A novel enzymic activity of phenylalanyl transfer ribonucleic acid synthetase from baker's yeast: zinc ion induced transfer ribonucleic acid independent hydrolysis of adenosine triphosphate

Igloi¹,

Haar²,

Cramer³

1980

Biochemistry

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“…The amino group was critical for function and could not be successfully replaced by hydrogen [in 3-(4-imidazolyl)-l-propanol] or by chloro or bromo groups [as in 3-(4-imidazolyl)-2-chloro-1propanol]. The latter compounds, which have been employed as active-site-directed modifiers of LADH (Dahl et al, 1979), were synthesized with the hope of generating suicide substrates for HDH; however none of the alcohols, the acids, or the methyl esters delectably inactivated the enzyme. When the amino group was replaced by a hydroxyl, poor substrate activity was observed, suggesting a key role for this amino group in catalysis.…”

Section: Discussionmentioning

confidence: 99%

Salmonella typhimurium histidinol dehydrogenase: complete reaction stereochemistry and active site mapping

Grubmeyer¹,

Insinga²,

Bhatia³

et al. 1989

Biochemistry

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The stereochemistry of the L-histidinol dehydrogenase reaction was determined to be R at NAD for both steps, confirming previous results with a fungal extract [Davies, D., Teixeira, A., & Kenworthy, P. (1972) Biochem. J. 127, 335-343]. NMR analysis of monodeuteriohistidinols produced by histidinol/NADH exchange reactions arising via reversal of the alcohol oxidation reaction indicated a single stereochemistry at histidinol for that step. Comparison of vicinal coupling values of the exchange products with those of L-alaninol and a series of (S)-2-amino-1-alcohols allowed identification of the absolute stereochemistry of monodeuteriohistidinols and showed that histidinol dehydrogenase removes first the pro-S then the pro-R hydrogens of substrate histidinol. The enzyme stereochemistry was confirmed by isotope effects for monodeuteriohistidinols as substrates for the pro-R-specific dehydrogenation catalyzed by liver alcohol dehydrogenase. Active site mapping was undertaken to investigate substrate-protein interactions elsewhere in the histidinol binding site. Critical binding regions are the side-chain amino group and the imidazole ring, whose methylation at the 1- or 2-position caused severe decreases in binding affinity. Use of alternative substrates further clarified active site interactions with the substrate. Compounds in which the alpha-amino group was replaced by chloro, bromo, or hydrogen substituents were not substrates of the overall reaction at 1/10,000 the normal rate.(ABSTRACT TRUNCATED AT 250 WORDS)

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“…However, work is in progress to determine the kinetic constants of inactivation of SDH by the different enantiomers of the chloro analogues, D and ~-a-chloro-~-(5-imidazolyl)propionic acid, as was previously determined for horse liver ADH [28].…”

Section: The Mechanism For the Labelling Of Sdh With Brimppohmentioning

confidence: 99%

Affinity labelling of sorbitol dehydrogenase from sheep liver with α‐bromo‐β‐(5‐imidazolyl)propionic acid

Reiersen

Sletten

McKinley-McKee

1993

European Journal of Biochemistry

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The metal-directed alkylating agent ~~-a-bromo-~-(5-imidazolyl)propionic acid (BrImPpOH) is shown to be an affinity-labelling reagent for sheep liver sorbitol dehydrogenase (SDH). As previously found for horse liver alcohol dehydrogenase (ADH), it modifies a cysteine ligand to the active-site zinc. In this case it is selectively incorporated (over 90%) at Cys43 in each of the four polypeptide chains/protomers of sheep liver SDH. Incorporated reagent and residual activity correlated. The first order inactivation constant, k,, and KEI, the dissociation constant for SDH and BrImPpOH, have been determined at different pH. The reactivity of BrImPpOH for SDH is higher than that for horse liver and yeast ADH. The protection of SDH against BrImPpOH inactivation by buffers and other molecules shows some similarities to that with horse liver ADH. However, sheep liver SDH bound BrImPpOH, imidazole and phosphate ions much weaker than liver ADH. The pK, values from the plot of log(k*/&I) against pH are approximately 7.0 and 8.8-8.9. The former pK, value probably represents ionization of an imidazole group and the latter the zindwater ionization in SDH. These pK, values are similar to those found for horse liver ADH. They are apparently not noticeably influenced by a second cysteine ligand in liver ADH being replaced by a proposed glutamic acid residue as a ligand to the catalytic zinc in SDH. The plot of logk, against pH shows pK, values around 7.0 and 9.2 for the SDH-BrImPpOH-complex. The pK, of 7.0 is the same as for log(kJK,,), and indicates no significant perturbation due to the binding of BrImPpOH to SDH. The pK, around 9.2 indicates perturbation of the zindwater ionization or the ionization of Cys43.Sorbitol dehydrogenase (SDH) is present at different tissues [l] and is an important marker for damage in the liver and testis [2-41. SDH is involved in the sorbitol or polyol pathway, an important alternative route for glucose metabolism to fructose via sorbitol by first aldose reductase then sorbitol dehydrogenase [ 11. SDH and alcohol dehydrogenase (ADH) are related through their sequences [5 -101. Sequence similarity of sheep liver SDH and the long-chain zinc ADH indicates that there are close structural similarities between them [7,11, 121. The sheep liver SDH sequence has recently been corrected in the order of N-terminal positions 2-5 and by the addition of an extra histidine residue in position 52 [9]. SDH is a tetramer (152 kDa) of identical subunits like yeast ADH. Unlike liver ADH which is a dimer of identical subunits each containing a catalytic and a structural zinc atom, SDH has only 1 zinc atodsubunit [13-151. This is a catalytic zinc atom, as indicated by inhibition experiments [ 14 -201. Enzymes. Sorbitol dehydrogenase, L-iditol : NAD' 2-oxidoreductase (EC 1.1.1.14); dcohol dehydrogenase, alcohol: NAD' oxidoreductase (EC 1.1.1.1); trypsin (EC 3.4.21.4); thermolysin (EC 3.4.24.4); endoproteinase Asp-N (EC 3.4.24.33).A computer model of the sheep liver SDH protomer based on its sequence (355 residues) and th...

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Metal‐directed affinity labelling

Cited by 13 publications

References 12 publications

A novel enzymic activity of phenylalanyl transfer ribonucleic acid synthetase from baker's yeast: zinc ion induced transfer ribonucleic acid independent hydrolysis of adenosine triphosphate

A novel enzymic activity of phenylalanyl transfer ribonucleic acid synthetase from baker's yeast: zinc ion induced transfer ribonucleic acid independent hydrolysis of adenosine triphosphate

Salmonella typhimurium histidinol dehydrogenase: complete reaction stereochemistry and active site mapping

Affinity labelling of sorbitol dehydrogenase from sheep liver with α‐bromo‐β‐(5‐imidazolyl)propionic acid

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