title: Expression of a hyperthermophilic aspartate aminotransferase in Escherichia coli.
abstract: The gene for an archaebacterial hyperthermophilic enzyme, aspartate aminotransferase from Sulfolobus solfataricus (AspATSs), was expressed in Escherichia coli and the enzyme purified to homogeneity. A suitable expression vector and host strain were selected and culture conditions were optimized so that 6-7 mg of pure enzyme per litre of culture were obtained repeatedly. The recombinant enzyme and the authentic AspATSs are indistinguishable: in fact, they have the same molecular weight, estimated by means of SDS-PAGE and gel filtration, the same Km values for 2-oxo-glutarate and cysteine sulphinate and the same UV-visible spectra. Moreover, recombinant AspATSs is thermophilic and thermostable just as the enzyme extracted from Sulfolobus solfataricus. The protocol described may be used to produce thermostable arachaebacterial enzymes in mesophilic hosts.
authors.affiliation: Dipartimento di Chimica Organica e Biologica, Università di Napoli, Italy.
authors.forename: M I
authors.initials: MI
authors.lastname: Arnone
authors.forename: L
authors.initials: L
authors.lastname: Birolo
authors.forename: M V
authors.initials: MV
authors.lastname: Cubellis
authors.forename: G
authors.initials: G
authors.lastname: Nitti
authors.forename: G
authors.initials: G
authors.lastname: Marino
authors.forename: G
authors.initials: G
authors.lastname: Sannia
mesh_terms: D001219:Aspartate Aminotransferases / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D004926:Escherichia coli / Q000201:enzymology*; D008969:Molecular Sequence Data; D009841:Oligonucleotides; D010957:Plasmids; D011994:Recombinant Proteins / Q000302:isolation & purification; D016944:Sulfolobus / Q000201:enzymology
chemical_list: D009841:Oligonucleotides; D011994:Recombinant Proteins; D001219:Aspartate Aminotransferases