The priB and priC replication proteins of Escherichia coli. Genes, DNA sequence, overexpression, and purification.
The Escherichia coli DNA replication proteins n and n" function in vitro in the assembly of the primosome, a mobile multiprotein replication priming complex thought to operate on the lagging-strand template at the E. coli DNA replication fork. Both proteins have been purified from E. coli HMS83 cells based on their requirement for the reconstitution of bacteriophage phi X174 complementary strand DNA synthesis in vitro with purified proteins. As a step toward understanding the role of these proteins in vivo, the genes for primosomal proteins n and n", designated priB and priC, respectively, have been cloned molecularly. priB encodes a 104-amino acid 11.4-kDa polypeptide and corresponds to an previously identified open reading frame between rpsF and rps R within a ribosomal protein operon at 95.5 min on the E. coli chromosome. priC encodes a 175-amino acid 20.3-kDa polypeptide. These two gene products were overexpressed at least 1000-fold in E. coli using a bacteriophage T7 transient expression system. Both proteins have been purified to apparent homogeneity from extracts prepared from these overproducing strains.
Zavitz
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KH
Program in Molecular Biology, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
DiGate
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Marians
K J
KJ
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular; D003062:Codon; D004261:DNA Replication; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015870:Gene Expression; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000302:isolation & purification
D001426:Bacterial Proteins; D003062:Codon; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; C069996:PriB protein (N-terminal); C069995:PriC protein, E coli