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Probing enzymic transition state hydrophobicities

P P Wangikar1, J O Rich, D S Clark

  • 1Department of Chemical and Biochemical Engineering, University of Iowa, Iowa City 52242, USA.

Biochemistry
|September 26, 1995
PubMed
Summary

Hydrophobic interactions in enzymes are crucial, even in nonaqueous solvents. Protein engineering and solvent changes can modify these interactions, enabling tailored enzyme function.

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Area of Science:

  • Biochemistry
  • Enzymology
  • Protein Engineering

Background:

  • Hydrophobic interactions are vital in biological processes but poorly understood in nonaqueous enzymology.
  • Understanding these interactions is key to controlling enzyme function in diverse media.

Purpose of the Study:

  • To estimate enzyme-substrate hydrophobic interactions for subtilisin BPN' in various solvents.
  • To develop a method for quantifying enzymic transition state hydrophobicity.
  • To investigate the impact of protein engineering on binding pocket hydrophobicity.

Main Methods:

  • Utilized a model reaction catalyzed by subtilisin BPN' in different solvents.
  • Employed protein engineering by mutating the Gly166 residue to alter binding pocket hydrophobicity.
  • Developed a method to estimate transition state hydrophobicity by varying substrate and solvent properties.

Main Results:

  • Enzyme-substrate hydrophobic interactions and transition state stabilization are present in hydrophobic organic solvents.
  • Mutating Gly166 to alanine (G166A) and valine (G166V) increased S1 binding pocket hydrophobicity.
  • The developed method for calculating binding pocket hydrophobicity is applicable to other enzymes like horseradish peroxidase and alpha-chymotrypsin.

Conclusions:

  • Hydrophobic interactions significantly influence enzyme catalysis in both aqueous and nonaqueous environments.
  • Protein engineering offers a route to modulate enzyme function by altering binding pocket hydrophobicity.
  • The developed method provides a valuable tool for characterizing and tailoring enzyme activity.

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