Control of enzyme activity by an engineered disulfide bond.
A novel approach to the control of enzyme catalysis is presented in which a disulfide bond engineered into the active-site cleft of bacteriophage T4 lysozyme is capable of switching the activity on and off. Two cysteines (Thr21----Cys and Thr142----Cys) were introduced by oligonucleotide-directed mutagenesis into the active-site cleft. These cysteines spontaneously formed a disulfide bond under oxidative conditions in vitro, and the catalytic activity of the oxidized (cross-linked) T4 lysozyme was completely lost. On exposure to reducing agent, however, the disulfide bond was rapidly broken, and the reduced (non-cross-linked) lysozyme was restored to full activity. Thus an enzyme has been engineered such that redox potential can be used to control catalytic activity.
Matsumura
M
M
Institute of Molecular Biology, University of Oregon, Eugene 97403.
Matthews
B W
BW
D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D004252:DNA Mutational Analysis; D004220:Disulfides; D008958:Models, Molecular; D009113:Muramidase / Q000502:physiology*; D015202:Protein Engineering; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship; D013604:T-Phages / Q000201:enzymology
D004220:Disulfides; D011994:Recombinant Proteins; D009113:Muramidase