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The use of genetically engineered tryptophan to identify the movement of a domain of B. stearothermophilus lactate dehydrogenase with the process which limits the steady-state turnover of the enzyme.

Authors
  • Waldman, A D
  • Hart, K W
  • Clarke, A R
  • Wigley, D B
  • Barstow, D A
  • Atkinson, T
  • Chia, W N
  • Holbrook, J J
Type
Published Article
Journal
Biochemical and biophysical research communications
Publication Date
Jan 29, 1988
Volume
150
Issue
2
Pages
752–759
Identifiers
PMID: 3422557
Source
Medline
License
Unknown

Abstract

A general technique for monitoring the intramolecular motion of a protein is described. Genetic engineering is used to replace all the natural tryptophan residues with tyrosine. A single tryptophan residue is then inserted at a specific site within the protein where motion is then detected from the fluorescence characteristics of this fluorophore. This technique has been used in B. stearothermophilus lactate dehydrogenase mutant (W80Y, W150Y, W203Y, G106W) to correlate the slow closure of a surface loop of polypeptide (residues 98-110) with the maximum catalytic velocity of the enzyme.

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