The enthalpimetric determination of inhibition constants for the inhibition of urease by acetohydroxamic acid

• Autorzy: Zaborska W. , Leszko M. , Kot M. , Juszkiewicz A.
• Języki publikacji: EN

Abstrakty

EN

The effect of concentration of acetohydroxamic acid (AHA) on inhibition of jack bean urease in phosphate buffer, pH 7.0, at 25°C, was studied. The meas­urements were performed at urease concentration of 2.5 mg/100 cm3 for con­centrations of urea and AHA ranging in the range of 2-50 mmol dm-3 and 0.25-10 mmol dm-3, respectively. The reactions were monitored by two techniques: analytical and enthalpimetric. For the analytical technique the growth of ammonia concentration in the course of the reaction was determined. From the recorded progress curves the following parameters were calculated for each inhibitor concentration: the initial reaction rate, the steady-state rate and the inversion constant. From these parameters the inhibition constants of the initial and steady-state stages of the reaction, K\ and K\y were calculated. The former constant did not change whereas the latter one proved to decrease quickly with an increase in inhibitor concentration. This behaviour resulted from the fact that the inactive complex EI* was not a product of internal inversion but was formed in the reaction: — 2/3I + EI —> (EI • — 2/3I)*. The disso3ciation constant of this complex is equal to about 0.3 x 10-3 (mol dm-3)2/3.

Słowa kluczowe

EN

inhibition constant; urease concentration; urease inhibition; acetohydroxamic acid; enzyme calorimetry

• Wydawca: -
• Czasopismo: Acta Biochimica Polonica
• Rocznik: 1997
• Tom: 44
• Numer: 1
• Opis fizyczny: p.89-98,fig.

Twórcy

autor

Zaborska W.

• Jagiellonian University, R.Ingardena 3, 30-060 Cracow, Poland

autor

Leszko M.

autor

Kot M.

autor

Juszkiewicz A.

Bibliografia

• 1. Kobashi, K., Hase, J.-I. & Uehara, K. (1962) Specific inhibition of urease by hydroxamic acids. Biochim. Biophys. Acta 65, 380-383.
• 2. Kobashi, K., Kumakd, K & Hase, J.-I. (1971) Effect of acyl residues of hydroxamic acids on urease inhibition. Biochim. Biophys. Acta 227, 429-441.
• 3. Kobashi, K & Hase, J.-I. (1967) Inhibition of Proteus vulgaris urease by hydroxamic acids. J. Biochem. (Tokyo) 62, 293-299.
• 4. Mobley, H.L.T. & Hausinger, R.P. (1989) Mi­crobial ureases: Significance, regulation, and molecular characterization. Microbiol. Rev. 53, 85-108.
• 5. Blakeley, R.L., Webb, E.C. & Zerner, B. (1969) Jack bean urease: A new purification proce­dure and reliable rate assay. Biochemistry 8, 1984-1990.
• 6. Dixon, N.E., Hinds, J.A., Fihelly, A.K., Gaz- • zola, C., Winzor, D.J., Blakeley, R.L. & Zer- ner, B. (1980) Jack bean urease (EC 3.5.1.5). IV. The molecular size and the mechanism of inhibition by hydroxamic acids. Spectrophoto- metric titration of enzymes with reversible inhibitors. Can. J. Biochem. 58,1323-1334.
• 7. Dixon, N.E., Riddles, P.W., Gazzola, C. & Zerner, B. (1980) Jack bean urease (EC 3.5.1.5.). V. On the mechanism of action of urease on urea, formamide, acetamide, N- mcthylurca and related compounds. Can. J. Biochem. 58,1335-1344.
• 8. Blakeley, R.L. & Zerner, B. (1984) Jack bean urease: the first nickel enzyme. J. Mol. Ca­talysis 23, 263-292.
• 9. Todd, M.J. & Hausinger, R.P. (1989) Competi­tive inhibitors of Klebsiella aerogenes urease. J. Biol. Chem. 264, 15835-15842.
• 10. Morrison, J.F. & Walsh, C.T. (1988) The be­haviour and significance of slow-binding en­zyme inhibitors. Adv. Enzymol. 61, 201- 301.
• 11. Krajewska, B., Leszko, M. & Zaborska, W. (1990) Urease immobilized on chitosan mem­brane. Preparation and properties. J. Chem. Tech. Biotechnol. 48, 337-350.
• 12. Juszkiewicz, A., Kot, M., Leszko, M. & Zabor­ska, W. (1995) The enthalpimetric determina­tion of inhibition of urease by fluoride ion. Thermochim. Acta 249, 301-311.
• 13. Weatherbun, M.W. (1967) Phenol-hypochlo- ride reaction for determination of ammonia. Anal. Chem. 39, 971-974.
• 14. Witwicki, J., Ardclt, W. Elementy enzymolo- gii. PWN Warszawa, 1989 (in Polish).
• 15. Wilkinson, G.N. (1961) Statistical estima­tions in enzyme kinetics. Biochem. J. 80, 324-332.
• 16. Zaborska, W. & Leszko, M. (1994) Inhibition of activity of urease by phosphate buffer. Pol­ish J. Chem. 68, 2733-2739.
• 17. Olech, Z.I. (1993) Electron paramagnetic reso­nance spectrum of crystalline jack bean urease. Colloids Surfaces A: Physicochem. Eng. Aspects 72, 365-367.