Response of rat liver glutaminase to pH. Mediation by phosphate and ammonium ions

L. I. Szweda, D. E. Atkinson

Research output: Contribution to journalArticle

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Abstract

The activity of rat liver glutaminase from sedimented fractions of freeze-thawed mitochondria is strongly affected by variation in pH over a physiologically relevant range at approximate physiological concentrations of activators. As pH increases from 7.1 to 7.7 at 0.7 mM ammonium and 10 mM phosphate, the S0.5 for glutamine decreases 3.5-fold, from 38 to 11 mM. This results in an 8-fold increase in reaction velocity at 10 mM glutamine. In addition, the M0.5 for phosphate activation decreases from 21 to 8.9 mM as pH increases from 7.1 to 7.7. This apparent effect of pH on the affinity of glutaminase for phosphate is similar to previous reports of the pH effect on activation by ammonium (Verhoeven, A.J., Van Iwaarden, J.F., Joseph, S.K., and Meijer, A.J. (1983) Eur. J. Biochem. 133, 241-244; McGivan, J.D., and Bradford, N.M. (1988) Biochim. Biophys. Acta 159, 296-302). Glutaminase does not respond to variation in pH between 7.1 and 7.7 when phosphate and ammonium are saturating. The effects of the two modifiers are additive. Each is still effective, as is pH, when the other is saturating. Therefore, it appears that the effects of pH on the apparent affinity of the enzyme for ammonium and phosphate account for the enzyme's response to pH. These results may help explain previous reports of minimal effects of pH on glutaminase at saturating concentrations of related substances (McGivan, J.D., Lacey, J.H., and Joseph, K. (1980) Biochim. J. 192, 537-542; Horowitz, M.L., and Knox, W.E. (1968) Enzymol. Biol. Clin. 9, 241l-255; McGivan J.D., and Bradford, N.M. (1983) Biochim. Biophys. Acta 759, 296-302). Glutaminase binds glutamine cooperatively with Hill coefficients ranging from 1.7 to 2.2, which suggests at least two and probably three or more interacting binding sites for glutamine. The strong response of liver glutaminase to pH and the fact that the reaction can supply metabolites for urea synthesis suggest a possible regulatory role of glutaminase in ureagenesis.

Original languageEnglish (US)
Pages (from-to)15357-15360
Number of pages4
JournalJournal of Biological Chemistry
Volume264
Issue number26
StatePublished - Jan 1 1989

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Glutaminase
Liver
Rats
Ions
Glutamine
Phosphates
Ammonium Compounds
Chemical activation
pH effects
Mitochondria
Enzymes
Metabolites
ammonium phosphate
Urea
Binding Sites

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Response of rat liver glutaminase to pH. Mediation by phosphate and ammonium ions. / Szweda, L. I.; Atkinson, D. E.

In: Journal of Biological Chemistry, Vol. 264, No. 26, 01.01.1989, p. 15357-15360.

Research output: Contribution to journalArticle

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abstract = "The activity of rat liver glutaminase from sedimented fractions of freeze-thawed mitochondria is strongly affected by variation in pH over a physiologically relevant range at approximate physiological concentrations of activators. As pH increases from 7.1 to 7.7 at 0.7 mM ammonium and 10 mM phosphate, the S0.5 for glutamine decreases 3.5-fold, from 38 to 11 mM. This results in an 8-fold increase in reaction velocity at 10 mM glutamine. In addition, the M0.5 for phosphate activation decreases from 21 to 8.9 mM as pH increases from 7.1 to 7.7. This apparent effect of pH on the affinity of glutaminase for phosphate is similar to previous reports of the pH effect on activation by ammonium (Verhoeven, A.J., Van Iwaarden, J.F., Joseph, S.K., and Meijer, A.J. (1983) Eur. J. Biochem. 133, 241-244; McGivan, J.D., and Bradford, N.M. (1988) Biochim. Biophys. Acta 159, 296-302). Glutaminase does not respond to variation in pH between 7.1 and 7.7 when phosphate and ammonium are saturating. The effects of the two modifiers are additive. Each is still effective, as is pH, when the other is saturating. Therefore, it appears that the effects of pH on the apparent affinity of the enzyme for ammonium and phosphate account for the enzyme's response to pH. These results may help explain previous reports of minimal effects of pH on glutaminase at saturating concentrations of related substances (McGivan, J.D., Lacey, J.H., and Joseph, K. (1980) Biochim. J. 192, 537-542; Horowitz, M.L., and Knox, W.E. (1968) Enzymol. Biol. Clin. 9, 241l-255; McGivan J.D., and Bradford, N.M. (1983) Biochim. Biophys. Acta 759, 296-302). Glutaminase binds glutamine cooperatively with Hill coefficients ranging from 1.7 to 2.2, which suggests at least two and probably three or more interacting binding sites for glutamine. The strong response of liver glutaminase to pH and the fact that the reaction can supply metabolites for urea synthesis suggest a possible regulatory role of glutaminase in ureagenesis.",
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N2 - The activity of rat liver glutaminase from sedimented fractions of freeze-thawed mitochondria is strongly affected by variation in pH over a physiologically relevant range at approximate physiological concentrations of activators. As pH increases from 7.1 to 7.7 at 0.7 mM ammonium and 10 mM phosphate, the S0.5 for glutamine decreases 3.5-fold, from 38 to 11 mM. This results in an 8-fold increase in reaction velocity at 10 mM glutamine. In addition, the M0.5 for phosphate activation decreases from 21 to 8.9 mM as pH increases from 7.1 to 7.7. This apparent effect of pH on the affinity of glutaminase for phosphate is similar to previous reports of the pH effect on activation by ammonium (Verhoeven, A.J., Van Iwaarden, J.F., Joseph, S.K., and Meijer, A.J. (1983) Eur. J. Biochem. 133, 241-244; McGivan, J.D., and Bradford, N.M. (1988) Biochim. Biophys. Acta 159, 296-302). Glutaminase does not respond to variation in pH between 7.1 and 7.7 when phosphate and ammonium are saturating. The effects of the two modifiers are additive. Each is still effective, as is pH, when the other is saturating. Therefore, it appears that the effects of pH on the apparent affinity of the enzyme for ammonium and phosphate account for the enzyme's response to pH. These results may help explain previous reports of minimal effects of pH on glutaminase at saturating concentrations of related substances (McGivan, J.D., Lacey, J.H., and Joseph, K. (1980) Biochim. J. 192, 537-542; Horowitz, M.L., and Knox, W.E. (1968) Enzymol. Biol. Clin. 9, 241l-255; McGivan J.D., and Bradford, N.M. (1983) Biochim. Biophys. Acta 759, 296-302). Glutaminase binds glutamine cooperatively with Hill coefficients ranging from 1.7 to 2.2, which suggests at least two and probably three or more interacting binding sites for glutamine. The strong response of liver glutaminase to pH and the fact that the reaction can supply metabolites for urea synthesis suggest a possible regulatory role of glutaminase in ureagenesis.

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