Escherichia coli signal peptidase recognizes and cleaves archaeal signal sequence

Abstract

Tk1884, an open reading frame encoding α-amylase in Thermococcus kodakarensis, was cloned with the native signal sequence and expressed in Escherichia coli. Heterologous gene expression resulted in secretion of the recombinant protein to the extracellular culture medium. Extracellular α-amylase activity gradually increased after induction. Tk1884 was purified from the extracellular medium, and its molecular mass determined by electrospray ionization mass spectrometry indicated the cleavage of a few amino acids. The N-terminal amino acid sequence of the purified Tk1884 was determined, which revealed that the signal peptide was cleaved between Ala26 and Ala27 by E. coli signal peptidase. To the best of our knowledge, this is the first report describing an archaeal signal sequence recognized and cleaved by E. coli signal peptidase.

Complete signal peptide of Tk1884, an α-amylase from Thermococcus kodakarensis, is not necessary for extracellular secretion of the enzyme by Escherichia coli

“…E. coli signal peptidase cleaved the signal peptide consisting of 26 amino acids [22]. Here we demonstrate the effect of truncation of signal peptide on the secretion of Tk1884.…”

“…We have previously produced Tk1884, an α-amylase from hyperthermophilic archaeon T. kodakarensis, with the native signal sequence in E. coli and demonstrated that signal peptidase of the host cleaved the signal sequence of the archaeal origin and the mature protein was secreted in the extracellular medium [22]. In the present study we have shown that deletion of the sixteen N-terminal amino acids, out of twenty six, of the signal peptide resulted in the secretion of the recombinant protein to the extracellular medium but without cleavage of the signal peptide.…”

Escherichia coli Signal Peptidases Cleave the Signal Sequence of TK0522, a Carbohydrate Esterase from Hyperthermophilic Archaeon Thermococcus kodakarensis

“…There have been reports on production and extracellular secretion of recombinant bacterial enzymes produced in E. coli, including α-amylase (Shahhoseini, 2003;Yamabhai et al, 2008;Malik et al, 2013), chitinase (Yamabhai et al, 2008); mannanase (Zhang et al, 2006), subtilisin (Ikemura et al, 1987) and xylanase (Jalal et al, 2011). Apart from bacterial enzymes, production and secretion of archaeal enzymes, using their native signal peptide, have also been reported in E. coli (Muhammad et al, 2017). We have previously cloned the TK0522 gene encoding the mature carbohydrate esterase from Thermococcus kodakarensis and characterized the recombinant protein (Tariq et al, 2019).…”