Development of β-Lactamase as a Tool for Monitoring Conditional Gene Expression by a Tetracycline-Riboswitch in Methanosarcina acetivorans
Figure 1
Synthesis of β-lactamase in M. acetivorans. (a) scheme of blaN (bla gene encoding β-lactamase and lacking the sequence for signal peptide) fusions inserted into the host chromosome; fsp: sequence encoding putative methanoarchaeal flagellin signal peptide; mcrB P: methanoarchaeal mcrB promoter; RBS: ribosome binding sequence; sps: signal peptide encoding sequence. (b) anaerobic cleavage of nitrocefin in cultures (C), culture supernatants (S), and cell lysates (E) of M. acetivorans strains carrying bla without (pBlaN) or with (pBlaNFSP) a putative archaeal signal peptide sequence on the chromosome, or the E. coli ampicillin resistance cassette on a self-replicating plasmid (pWM321), respectively; shown are sections of the culture tubes after 20 min of incubation at room temperature in the presence of 20 μM nitrocefin.
