title: Cooperative working of bacterial chromosome replication proteins generated by a reconstituted protein expression system.
abstract: Replication of all living cells relies on the multirounds flow of the central dogma. Especially, expression of DNA replication proteins is a key step to circulate the processes of the central dogma. Here we achieved the entire sequential transcription-translation-replication process by autonomous expression of chromosomal DNA replication machineries from a reconstituted transcription-translation system (PURE system). We found that low temperature is essential to express a complex protein, DNA polymerase III, in a single tube using the PURE system. Addition of the 13 genes, encoding initiator, DNA helicase, helicase loader, RNA primase and DNA polymerase III to the PURE system gave rise to a DNA replication system by a coupling manner. An artificial genetic circuit demonstrated that the DNA produced as a result of the replication is able to provide genetic information for proteins, indicating the in vitro central dogma can sequentially undergo two rounds.
authors.affiliation: Department of Bioengineering and Robotics, Tohoku University, 6-6-01, Aramakiaza-aoba, Aoba-ku, Sendai, Miyagi, 980-8579, Japan. fujiwara@molbot.mech.tohoku.ac.jp
authors.forename: Kei
authors.initials: K
authors.lastname: Fujiwara
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Katayama
authors.forename: Shin-Ichiro M
authors.initials: SM
authors.lastname: Nomura
mesh_terms: D002474:Cell-Free System; D002876:Chromosomes, Bacterial / Q000378:metabolism*; D004258:DNA Polymerase III / Q000378:metabolism; D019915:DNA Primase / Q000096:biosynthesis / Q000378:metabolism; D004261:DNA Replication; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D053263:Gene Regulatory Networks; D014176:Protein Biosynthesis; D021122:Protein Subunits / Q000096:biosynthesis / Q000235:genetics; D014158:Transcription, Genetic
chemical_list: D029968:Escherichia coli Proteins; D021122:Protein Subunits; D019915:DNA Primase; D004258:DNA Polymerase III