PMID:9278153
| Citation |
Valentini, G, Mattevi, A, Barilla, D, Galizzi, A and Speranza, ML (1997) Recombinant pyruvate kinase type I from Escherichia coli: overproduction and revised C-terminus of the polypeptide. Biol. Chem. 378:719-21 |
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| Abstract |
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. |
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| Keywords |
Amino Acid Sequence; Bacterial Proteins/biosynthesis; Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Base Sequence; Cloning, Molecular; Escherichia coli/enzymology; Escherichia coli/genetics; Molecular Sequence Data; Polymerase Chain Reaction; Pyruvate Kinase/biosynthesis; Pyruvate Kinase/chemistry; Pyruvate Kinase/genetics |
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