title: Total chemical synthesis of a thermostable enzyme capable of polymerase chain reaction.
abstract: Polymerase chain reaction (PCR) has been a defining tool in modern biology. Towards realizing mirror-image PCR, we have designed and chemically synthesized a mutant version of the 352-residue thermostableSulfolobus solfataricusP2 DNA polymerase IV with l-amino acids and tested its PCR activity biochemically. To the best of our knowledge, this enzyme is the largest chemically synthesized protein reported to date. We show that with optimization of PCR conditions, the fully synthetic polymerase is capable of amplifying template sequences of up to 1.5 kb. The establishment of this synthetic route for chemically synthesizing DNA polymerase IV is a stepping stone towards building a d-enzyme system for mirror-image PCR, which may open up an avenue for the creation of many mirror-image molecular tools such as mirror-image systematic evolution of ligands by exponential enrichment.
authors.affiliation: School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Center for Synthetic and Systems Biology, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University, Beijing, China; Department of Chemistry, Tsinghua-Peking Joint Center for Life Sciences, Ministry of Education Key Laboratory of Bioorganic Phosphorus Chemistry and Chemical Biology, Tsinghua University, Beijing, China.
authors.forename: Weiliang
authors.initials: W
authors.lastname: Xu
authors.affiliation: School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Center for Synthetic and Systems Biology, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University , Beijing, China.
authors.forename: Wenjun
authors.initials: W
authors.lastname: Jiang
authors.affiliation: Department of Chemistry, Tsinghua-Peking Joint Center for Life Sciences, Ministry of Education Key Laboratory of Bioorganic Phosphorus Chemistry and Chemical Biology, Tsinghua University , Beijing, China.
authors.forename: Jiaxing
authors.initials: J
authors.lastname: Wang
authors.affiliation: School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Center for Synthetic and Systems Biology, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University , Beijing, China.
authors.forename: Linping
authors.initials: L
authors.lastname: Yu
authors.affiliation: School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Center for Synthetic and Systems Biology, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University , Beijing, China.
authors.forename: Ji
authors.initials: J
authors.lastname: Chen
authors.affiliation: School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Center for Synthetic and Systems Biology, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University , Beijing, China.
authors.forename: Xianyu
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Chemistry, Tsinghua-Peking Joint Center for Life Sciences, Ministry of Education Key Laboratory of Bioorganic Phosphorus Chemistry and Chemical Biology, Tsinghua University , Beijing, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Liu
authors.affiliation: School of Life Sciences, Tsinghua-Peking Joint Center for Life Sciences, Center for Synthetic and Systems Biology, Ministry of Education Key Laboratory of Bioinformatics, Tsinghua University , Beijing, China.
authors.forename: Ting F
authors.initials: TF
authors.lastname: Zhu
keywords: Sulfolobus solfataricus P2 DNA polymerase IV; native chemical ligation; polymerase chain reaction; protein chemical synthesis; thermostable