The role of protein dynamics in the evolution of new enzyme function.
Enzymes must be ordered to allow the stabilization of transition states by their active sites, yet dynamic enough to adopt alternative conformations suited to other steps in their catalytic cycles. The biophysical principles that determine how specific protein dynamics evolve and how remote mutations affect catalytic activity are poorly understood. Here we examine a 'molecular fossil record' that was recently obtained during the laboratory evolution of a phosphotriesterase from Pseudomonas diminuta to an arylesterase. Analysis of the structures and dynamics of nine protein variants along this trajectory, and three rationally designed variants, reveals cycles of structural destabilization and repair, evolutionary pressure to 'freeze out' unproductive motions and sampling of distinct conformations with specific catalytic properties in bi-functional intermediates. This work establishes that changes to the conformational landscapes of proteins are an essential aspect of molecular evolution and that change in function can be achieved through enrichment of preexisting conformational sub-states.
Research School of Chemistry, Australian National University, Canberra, Australia.
Eleanor
E
Campbell
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, Canada.|Department of Biochemistry, University of Cambridge, Cambridge, UK.
Miriam
M
Kaltenbach
Research School of Chemistry, Australian National University, Canberra, Australia.
Galen J
GJ
Correy
Research School of Chemistry, Australian National University, Canberra, Australia.
Paul D
PD
Carr
Biomedicine Discovery Institute, Monash University, Clayton, Victoria, Australia.|Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
Benjamin T
BT
Porebski
Research School of Chemistry, Australian National University, Canberra, Australia.
Emma K
EK
Livingstone
Research School of Chemistry, Australian National University, Canberra, Australia.
Livnat
L
Afriat-Jurnou
Biomedicine Discovery Institute, Monash University, Clayton, Victoria, Australia.|Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
Ashley M
http://orcid.org/0000-0003-2943-9044
AM
Buckle
Institut de Biologie Structurale, University Grenoble Alpes, Commissariat à l'Energie Atomique and Centre National de la Recherche Scientifique, Grenoble, France.
Martin
M
Weik
Department of Biochemistry, University of Cambridge, Cambridge, UK.
Florian
http://orcid.org/0000-0002-1367-6312
F
Hollfelder
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, Canada.
Nobuhiko
N
Tokuriki
Research School of Chemistry, Australian National University, Canberra, Australia.
Colin J
http://orcid.org/0000-0001-6150-3822
CJ
Jackson
D055162:Biocatalysis; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry / Q000378:metabolism*; D019143:Evolution, Molecular; D044345:Phosphoric Triester Hydrolases / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D011549:Pseudomonas / Q000201:enzymology*
D002265:Carboxylic Ester Hydrolases; C019734:arylesterase; D044345:Phosphoric Triester Hydrolases