Recombinant pyruvate kinase type I from Escherichia coli: Overproduction and revised C-terminus of the polypeptide

G Valentini, A Mattevi, D Barilla, A Galizzi, M L Speranza

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The gene encoding pyruvate kinase type I (PKI) of Escherichia coli was amplified by PCR, cloned and sequenced. The gene product was overexpressed in E.coli, using an inducible T7 RNA polymerase-based expression system. The transformed cells contained sixtyfold the enzyme activity of the reference cells and enabled the purification of 30 mg of highly active PKI from 1 liter of culture.

The gene sequence was determined and found to be different from the one previously reported, i.e., the T nucleotide at position 1351 was missing. This resulted in a downstream shift of the stop codon, thus the deduced polypeptide was 470 amino acids long instead of 462. In addition the twelve C-terminal amino acids of the former sequence were changed.

Original languageEnglish
Pages (from-to)719-721
Number of pages3
JournalJournal of Biological Chemistry
Issue number7
Publication statusPublished - Jul 1997


  • cloning
  • overexpression
  • sequence
  • DNA

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