Binding of the bovine basic pancreatic trypsin inhibitor (kunitz) to the 33,000 mr and 54,000 mr species of human urokinase

Thermodynamic study

Paolo Ascenzi, Gino Amiconi, Martino Bolognesi, Enea Menegatti, Mario Guarneri

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

The effect of pH and temperature on the apparent association equilibrium constant (Ka) for the binding of the bovine basic pancreatic trypsin inhibitor (BPTI, Kunitz inhibitor) to the 33,000 Mr and 54,000 Mr species of human urokinase (EC 3.4.21.31) has been investigated. Under all the experimental conditions, values of Ka for BPTI binding to the 33,000 Mr and 54,000 Mr species of human urokinase are identical. On lowering the pH from 9.5 to 4.5, values of Ka (at 21.0°C) for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) decrease thus reflecting the acidic pK-shift of the His-57 catalytic residue from 6.9, in the free enzyme, to 5.1, in the proteinase:inhibitor complex. At pH8.0, values of the apparent ther-modynamic parameters for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) are: K, = 4.9 × 104M-1, ΔG° = -6.3kcal/mol, and ΔS° = +37 entropy units (all at 21.0°C); and ΔH° = + 4.6 kcal/mol (temperature independent over the explored range, from 5.0°C to 45.0°C). Thermodynamics of BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) have been analyzed in parallel with those of related serine (pro)enzyme/Kazal-and /Kunitz-type inhibitor systems. Considering the known molecular models, the observed binding behaviour of BPTI to human urokinase (33,000 Mr and 54,000 Mr species) was related to the inferred stereochemistry of the proteinase/inhibitor contact region.

Original languageEnglish
Pages (from-to)51-55
Number of pages5
JournalJournal of Enzyme Inhibition and Medicinal Chemistry
Volume4
Issue number1
DOIs
Publication statusPublished - 1990

Fingerprint

Aprotinin
Urokinase-Type Plasminogen Activator
Thermodynamics
Peptide Hydrolases
Stereochemistry
Temperature
Molecular Models
Equilibrium constants
Entropy
Enzymes
Serine
Association reactions

Keywords

  • Bovine basic pancreatic trypsin inhibitor (Kunitz inhibitor, BPTI)
  • PH and temperature effects (on proteinase:inhibitor complex formation)
  • Proteinase:inhibitor complex formation
  • Thermodynamics (of proteinase:inhibitor complex formation)

ASJC Scopus subject areas

  • Drug Discovery
  • Pharmacology
  • Molecular Medicine
  • Biochemistry

Cite this

Binding of the bovine basic pancreatic trypsin inhibitor (kunitz) to the 33,000 mr and 54,000 mr species of human urokinase : Thermodynamic study. / Ascenzi, Paolo; Amiconi, Gino; Bolognesi, Martino; Menegatti, Enea; Guarneri, Mario.

In: Journal of Enzyme Inhibition and Medicinal Chemistry, Vol. 4, No. 1, 1990, p. 51-55.

Research output: Contribution to journalArticle

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abstract = "The effect of pH and temperature on the apparent association equilibrium constant (Ka) for the binding of the bovine basic pancreatic trypsin inhibitor (BPTI, Kunitz inhibitor) to the 33,000 Mr and 54,000 Mr species of human urokinase (EC 3.4.21.31) has been investigated. Under all the experimental conditions, values of Ka for BPTI binding to the 33,000 Mr and 54,000 Mr species of human urokinase are identical. On lowering the pH from 9.5 to 4.5, values of Ka (at 21.0°C) for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) decrease thus reflecting the acidic pK-shift of the His-57 catalytic residue from 6.9, in the free enzyme, to 5.1, in the proteinase:inhibitor complex. At pH8.0, values of the apparent ther-modynamic parameters for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) are: K, = 4.9 × 104M-1, ΔG° = -6.3kcal/mol, and ΔS° = +37 entropy units (all at 21.0°C); and ΔH° = + 4.6 kcal/mol (temperature independent over the explored range, from 5.0°C to 45.0°C). Thermodynamics of BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) have been analyzed in parallel with those of related serine (pro)enzyme/Kazal-and /Kunitz-type inhibitor systems. Considering the known molecular models, the observed binding behaviour of BPTI to human urokinase (33,000 Mr and 54,000 Mr species) was related to the inferred stereochemistry of the proteinase/inhibitor contact region.",
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AU - Guarneri, Mario

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N2 - The effect of pH and temperature on the apparent association equilibrium constant (Ka) for the binding of the bovine basic pancreatic trypsin inhibitor (BPTI, Kunitz inhibitor) to the 33,000 Mr and 54,000 Mr species of human urokinase (EC 3.4.21.31) has been investigated. Under all the experimental conditions, values of Ka for BPTI binding to the 33,000 Mr and 54,000 Mr species of human urokinase are identical. On lowering the pH from 9.5 to 4.5, values of Ka (at 21.0°C) for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) decrease thus reflecting the acidic pK-shift of the His-57 catalytic residue from 6.9, in the free enzyme, to 5.1, in the proteinase:inhibitor complex. At pH8.0, values of the apparent ther-modynamic parameters for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) are: K, = 4.9 × 104M-1, ΔG° = -6.3kcal/mol, and ΔS° = +37 entropy units (all at 21.0°C); and ΔH° = + 4.6 kcal/mol (temperature independent over the explored range, from 5.0°C to 45.0°C). Thermodynamics of BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) have been analyzed in parallel with those of related serine (pro)enzyme/Kazal-and /Kunitz-type inhibitor systems. Considering the known molecular models, the observed binding behaviour of BPTI to human urokinase (33,000 Mr and 54,000 Mr species) was related to the inferred stereochemistry of the proteinase/inhibitor contact region.

AB - The effect of pH and temperature on the apparent association equilibrium constant (Ka) for the binding of the bovine basic pancreatic trypsin inhibitor (BPTI, Kunitz inhibitor) to the 33,000 Mr and 54,000 Mr species of human urokinase (EC 3.4.21.31) has been investigated. Under all the experimental conditions, values of Ka for BPTI binding to the 33,000 Mr and 54,000 Mr species of human urokinase are identical. On lowering the pH from 9.5 to 4.5, values of Ka (at 21.0°C) for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) decrease thus reflecting the acidic pK-shift of the His-57 catalytic residue from 6.9, in the free enzyme, to 5.1, in the proteinase:inhibitor complex. At pH8.0, values of the apparent ther-modynamic parameters for BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) are: K, = 4.9 × 104M-1, ΔG° = -6.3kcal/mol, and ΔS° = +37 entropy units (all at 21.0°C); and ΔH° = + 4.6 kcal/mol (temperature independent over the explored range, from 5.0°C to 45.0°C). Thermodynamics of BPTI binding to human urokinase (33,000 Mr and 54,000 Mr species) have been analyzed in parallel with those of related serine (pro)enzyme/Kazal-and /Kunitz-type inhibitor systems. Considering the known molecular models, the observed binding behaviour of BPTI to human urokinase (33,000 Mr and 54,000 Mr species) was related to the inferred stereochemistry of the proteinase/inhibitor contact region.

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