TY - JOUR
T1 - A DNA computing-based genetic program for in vitro protein evolution via constrained pseudomodule shuffling
AU - Rose, John A.
AU - Takano, Mitsunori
AU - Hagiya, Masami
AU - Suyama, Akira
N1 - Funding Information:
The authors thank K. Sakamoto of the University of Tokyo for helpful discussions, and S. Suzuki for aid during figure preparation. Support provided by a JSPS Postdoctoral Fellowship and by JST CREST.
PY - 2003/6
Y1 - 2003/6
N2 - An in vitro domainal shuffling strategy for protein evolution was proposed in (J. Kolkman and W. Stemmer, Nat. Biotech. 19 (423) 2001). Due to backhybridization, however this method appears unlikely to be an efficient means of iteratively generating massive libraries of combinatorially shuffled genes. Recombination at the domain level (30-300 residues) also appears too coarse to support the evolution of proteins with substantially new folds. In this work, the module (10-25 residues long) and pseudomodule are adopted as the fundamental units of protein structure. Each protein is modelled as an N to C-terminal tour of a digraph composed of pseudomodules. An in vitro method based on PNA-mediated Whiplash PCR (PWPCR), RNA-protein fusion, and restriction-based recombination, XWPCR is then presented for evolving proteins with a high affinity for a given motif, subject to the constraint that each corresponds to a walk on the pseudomodule digraph of interest. Simulations predict that PWPCR is an efficient method of producing massive, shuffled gene libraries encoding for proteins as long as roughly 600 residues.
AB - An in vitro domainal shuffling strategy for protein evolution was proposed in (J. Kolkman and W. Stemmer, Nat. Biotech. 19 (423) 2001). Due to backhybridization, however this method appears unlikely to be an efficient means of iteratively generating massive libraries of combinatorially shuffled genes. Recombination at the domain level (30-300 residues) also appears too coarse to support the evolution of proteins with substantially new folds. In this work, the module (10-25 residues long) and pseudomodule are adopted as the fundamental units of protein structure. Each protein is modelled as an N to C-terminal tour of a digraph composed of pseudomodules. An in vitro method based on PNA-mediated Whiplash PCR (PWPCR), RNA-protein fusion, and restriction-based recombination, XWPCR is then presented for evolving proteins with a high affinity for a given motif, subject to the constraint that each corresponds to a walk on the pseudomodule digraph of interest. Simulations predict that PWPCR is an efficient method of producing massive, shuffled gene libraries encoding for proteins as long as roughly 600 residues.
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U2 - 10.1023/A:1023932912559
DO - 10.1023/A:1023932912559
M3 - Article
AN - SCOPUS:3543079726
SN - 1389-2576
VL - 4
SP - 139
EP - 152
JO - Genetic Programming and Evolvable Machines
JF - Genetic Programming and Evolvable Machines
IS - 2
ER -