Transition state stabilization of subtilisins in organic media

Zu‐Feng ‐F Xu, Rhett Affleck, Pramod Wangikar, Valerie Suzawa, Jonathan S. Dordick, Douglas S. Clark

Research output: Contribution to journalArticlepeer-review

58 Scopus citations


Electrostatic forces are among the stabilizing interactions that contribute to the high degree of enzyme‐transition state complementarity. The active‐site polarity, which can differ substaintially from that of water, is thus an important determinant of transition state stabilization. Here we pose the question of whether the rate of an enzymatic reaction proceeding through a charged transition state can be increased by increasing the active‐site polarity in an organic solvent. The active‐site polarity of subtilisin has been reduced by dehydration and suspension in a nonpolar solvent (tetrahydrofuran), and then increased by adding water to the solvent. Enhancing the local polarity substantially increasing the rate of catalysis, implicating polarity as an important factor in stabilizing the charged tetrahedral transition state. Studies with subtilisins whose active sites have been modified by site‐directed mutagenesis support the role of polarity in transition state stabilization. © 1994 John Wiley & Sons, Inc.

Original languageEnglish
Pages (from-to)515-520
Number of pages6
JournalBiotechnology and Bioengineering
Issue number6
StatePublished - 15 Mar 1994
Externally publishedYes


  • EPR spectroscopy
  • organic solvents
  • subtilisin
  • transition state stabilization


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