Expression of C5 protein, the protein component of Escherichia coli RNase P, from the tac promoter.
The accurate function of C5 protein, the protein component of Escherichia coli RNase P, is uncertain in vivo. A controllable expression system for C5 protein was constructed which can be used to investigate effects of C5 protein on various cellular functions including biosynthesis of RNase P in vivo. The semisynthetic rnpA gene encoding C5 protein was fused to the tac promoter of the pKK223-3 expression vector. This tac promoter expression system produced a high level of C5 protein upon induction with isopropyl-beta-D-thiogalacto-pyranoside. When the overexpressed C5 protein was purified and used for reconstitution of RNase P, the reconstituted enzyme was active. The N-terminal amino acid of the overexpressed C5 protein was leucine specified by the second codon of the rnpA gene. The more controllable expression system was constructed by introducing the lacIq gene into the vector sequence itself.
Park
B H
BH
Department of Chemistry, Korea Advanced Institute of Science and Technology, Taejon, Korea.
Choi
Y N
YN
Park
J W
JW
Sim
S
S
Gil
M C
MC
Kim
S
S
Kim
M
M
Lee
Y
Y
D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D001483:Base Sequence; D004722:Endoribonucleases / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000235:genetics*; D016337:RNA, Catalytic / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics; D043262:Ribonuclease P
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D016337:RNA, Catalytic; D007544:Isopropyl Thiogalactoside; D004722:Endoribonucleases; D043262:Ribonuclease P; C470865:ribonuclease P, E coli