Computational redesign of a mononuclear zinc metalloenzyme for organophosphate hydrolysis.
The ability to redesign enzymes to catalyze noncognate chemical transformations would have wide-ranging applications. We developed a computational method for repurposing the reactivity of metalloenzyme active site functional groups to catalyze new reactions. Using this method, we engineered a zinc-containing mouse adenosine deaminase to catalyze the hydrolysis of a model organophosphate with a catalytic efficiency (k(cat)/K(m)) of ~10(4) M(-1) s(-1) after directed evolution. In the high-resolution crystal structure of the enzyme, all but one of the designed residues adopt the designed conformation. The designed enzyme efficiently catalyzes the hydrolysis of the R(P) isomer of a coumarinyl analog of the nerve agent cyclosarin, and it shows marked substrate selectivity for coumarinyl leaving groups. Computational redesign of native enzyme active sites complements directed evolution methods and offers a general approach for exploring their untapped catalytic potential for new reactivities.
Department of Biochemistry, University of Washington, Seattle, Washington, USA.
Sagar D
SD
Khare
Yakov
Y
Kipnis
Per
P
Greisen
Ryo
R
Takeuchi
Yacov
Y
Ashani
Moshe
M
Goldsmith
Yifan
Y
Song
Jasmine L
JL
Gallaher
Israel
I
Silman
Haim
H
Leader
Joel L
JL
Sussman
Barry L
BL
Stoddard
Dan S
DS
Tawfik
David
D
Baker
D000243:Adenosine Deaminase / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D055162:Biocatalysis; D020134:Catalytic Domain; D019295:Computational Biology; D003198:Computer Simulation; D017076:Computer-Aided Design; D006868:Hydrolysis; D008667:Metalloproteins / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008958:Models, Molecular; D008968:Molecular Conformation; D009943:Organophosphorus Compounds / Q000737:chemistry / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry* / Q000378:metabolism
D008667:Metalloproteins; D009943:Organophosphorus Compounds; D000243:Adenosine Deaminase; D015032:Zinc