title: Cloning, Escherichia coli expression, and phase-transition chromatography-based purification of recombinant rabbit heart mitochondrial creatine kinase.
abstract: A cDNA clone of the mitochondrial sarcomeric creatine kinase cDNA was obtained by screening a rabbit heart library. This cDNA is characterized by a 1257-nucleotide open reading frame encoding a 419-amino-acid protein with a cleavable 39-amino-acid mitochondrial presequence (Accession No. AJ011334). This new member of the guanidino kinase family shows a high degree of sequence similarity with the other phosphagen kinases sequenced so far. The mature enzyme was efficiently expressed in Escherichia coli BL21(DE3) cells as a soluble octameric protein using the pET21 plasmid and purified by a three-step improved method including a final phase-transition chromatography.
authors.affiliation: UFR Chimie-Biochimie, Université Claude Bernard Lyon I, UPRESA CNRS 5013, 43 Boulevard du 11 Novembre 1918, Villeurbanne, F-69622, France.
authors.forename: O
authors.initials: O
authors.lastname: Marcillat
authors.forename: C
authors.initials: C
authors.lastname: Perraut
authors.forename: T
authors.initials: T
authors.lastname: Granjon
authors.forename: C
authors.initials: C
authors.lastname: Vial
authors.forename: M J
authors.initials: MJ
authors.lastname: Vacheron
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002845:Chromatography / Q000379:methods; D003001:Cloning, Molecular; D003402:Creatine Kinase / Q000235:genetics* / Q000302:isolation & purification*; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D007527:Isoenzymes; D008929:Mitochondria, Heart / Q000201:enzymology*; D008969:Molecular Sequence Data; D011817:Rabbits; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification
chemical_list: D018076:DNA, Complementary; D007527:Isoenzymes; D011994:Recombinant Proteins; D003402:Creatine Kinase