Importance of barrier shape in enzyme-catalyzed reactions. Vibrationally assisted hydrogen tunneling in tryptophan tryptophylquinone-dependent amine dehydrogenases

J Biol Chem. 2001 Mar 2;276(9):6234-42. doi: 10.1074/jbc.M008141200. Epub 2000 Nov 21.

Abstract

C-H bond breakage by tryptophan tryptophylquinone (TTQ)-dependent methylamine dehydrogenase (MADH) occurs by vibrationally assisted tunneling (Basran, J., Sutcliffe, M. J., and Scrutton, N. S. (1999) Biochemistry 38, 3218--3222). We show here a similar mechanism in TTQ-dependent aromatic amine dehydrogenase (AADH). The rate of TTQ reduction by dopamine in AADH has a large, temperature independent kinetic isotope effect (KIE = 12.9 +/- 0.2), which is highly suggestive of vibrationally assisted tunneling. H-transfer is compromised with benzylamine as substrate and the KIE is deflated (4.8 +/- 0.2). The KIE is temperature-independent, but reaction rates are strongly dependent on temperature. With tryptamine as substrate reaction rates can be determined only at low temperature as C-H bond cleavage is rapid, and an exceptionally large KIE (54.7 +/- 1.0) is observed. Studies with deuterated tryptamine suggest vibrationally assisted tunneling is the mechanism of deuterium and, by inference, hydrogen transfer. Bond cleavage by MADH using a slow substrate (ethanolamine) occurs with an inflated KIE (14.7 +/- 0.2 at 25 degrees C). The KIE is temperature-dependent, consistent with differential tunneling of protium and deuterium. Our observations illustrate the different modes of H-transfer in MADH and AADH with fast and slow substrates and highlight the importance of barrier shape in determining reaction rate.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Benzylamines / chemistry
  • Catalysis
  • Ethanolamine / metabolism
  • Indolequinones*
  • Oxidoreductases Acting on CH-NH Group Donors / chemistry*
  • Oxidoreductases Acting on CH-NH Group Donors / metabolism
  • Quinones / metabolism*
  • Tryptamines / chemistry
  • Tryptophan / analogs & derivatives*
  • Tryptophan / metabolism*
  • Vibration

Substances

  • Benzylamines
  • Indolequinones
  • Quinones
  • Tryptamines
  • tryptophan tryptophylquinone
  • tryptamine
  • Ethanolamine
  • Tryptophan
  • benzylamine
  • methylamine dehydrogenase
  • Oxidoreductases Acting on CH-NH Group Donors
  • aromatic amine dehydrogenase