Synergism in replication and translation of messenger RNA in a cell-free system.
Combination of the Q beta replicase reaction with the Escherichia coli cell-free translation system markedly enhances replication of a recombinant RQ-DHFR RNA consisting of the dihydrofolate reductase (DHFR) mRNA sequence inserted into RQ135(-1) RNA, an efficient naturally occurring Q beta replicase template. The enhancement is associated with a replication asymmetry previously described for the replication of Q beta phage RNA in vivo; the sense (+)-strands are produced in large excess over the antisense (-)-strands. This, in turn, results in increased synthesis of the functionally active DHFR. These effects are not observed when DHFR mRNAs or RQ135(-1) RNAs are used as templates, if the translation system is not complete, or if it is inhibited by puromycin. The coupled replication-translation of nonviral mRNA recombinants can serve as a useful model for studying the fundamental aspects of virus amplification and can be implemented for large-scale protein synthesis in vitro.
Morozov
I Y
IY
Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region.
Ugarov
V I
VI
Chetverin
A B
AB
Spirin
A S
AS
D001483:Base Sequence; D002474:Cell-Free System; D004274:DNA, Recombinant; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D014176:Protein Biosynthesis; D011777:Q beta Replicase / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics
D004274:DNA, Recombinant; D012333:RNA, Messenger; D012367:RNA, Viral; D013762:Tetrahydrofolate Dehydrogenase; D011777:Q beta Replicase