Recombinant Tk0522, a carbohydrate esterase homologue from Thermococcus kodakarensis, does not require a signal sequence for translocation to periplasmic space in Escherichia coli

We have cloned Tk0522, a carbohydrate esterase homologue from Thermococcus kodakarensis, with the deletion of a signal sequence of nineteen amino acids at the N-terminal and produced the recombinant Tk0522 in Escherichia coli. Recombinant Tk0522 was produced in soluble and enzymatically active form. Analysis of the cytosolic, periplasmic and extracellular fractions demonstrated the presence of Tk0522 in cytosolic and periplasmic fractions. The esterase activity in the periplasmic fraction, similar to the cytosolic fraction, increased with the passage of time after induction. No significant amount of esterase activity could be detected in the extracellular fraction. To the best of our knowledge, this is the first report on translocation of a mature archaeal esterase from cytosolic to periplasmic space in E. coli.