title: Expression, purification and characterisation of a novel mutant of the human protein kinase CK2.
abstract: The recombinant catalytic subunit of human protein kinase CK2 bas been mutagenised at the C-terminal region in an attempt to induce this tail to fold. We suppose in fact that this unstructured C-terminus just might be responsible for the high degradability of the human enzyme. On the basis of theoretical calculations we choose to substitute two distal prolines with alanines (PA 382-384). The mutant bas been purified to the electrophoretic homogeneity by means of three chromatographic steps. By circular dichroism Spectroscopy we verified if the double amino acids substitution reflected on the secondary structure of the recombinant a subunit. According to our theoretical predictions, we observed that the alpha-helix content of the protein increased when the two distal prolines were substituted by alanines. Moreover the mutant catalytic subunit shows a reduced ability to bind a classical inhibitor such as heparin.
authors.affiliation: Department of Biophysical Sciences and Technologies M.&O., School of Medicine, University of Genova, Genova, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Grasselli
authors.forename: Graziano
authors.initials: G
authors.lastname: Noviello
authors.forename: Cristina
authors.initials: C
authors.lastname: Rando
authors.forename: Claudio
authors.initials: C
authors.lastname: Nicolini
authors.forename: Laura
authors.initials: L
authors.lastname: Vergani
mesh_terms: D000595:Amino Acid Sequence; D047390:Casein Kinase II; D002942:Circular Dichroism; D003001:Cloning, Molecular; D015870:Gene Expression; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis; D016415:Sequence Alignment
chemical_list: D011994:Recombinant Proteins; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases