title: Cloning and purification of protein kinase CK2 recombinant alpha and beta subunits from the Mediterranean fly Ceratitis capitata.
abstract: The Mediterranean fruit fly Ceratitis capitata is an insect capable of wreaking extensive damage to a wide range of fruit crops. Protein kinase CK2 is a ubiquitous Ser/Thr kinase that is highly conserved among eukaryotes; it is a heterotetramer composed of two catalytic (α) and a dimer of regulatory (β) subunits. We present here the construction of the cDNA molecules of the CK2α and CK2β subunits from the medfly C. capitata by the 5'/3' RACE and RT-PCR methods, respectively. CcCK2α catalytic subunit presents the characteristic and conserved features of a typical protein kinase, similar to the regulatory CcCK2β subunit, that also possess the conserved features of regulatory CK2β subunits, as revealed by comparison of their predicted amino acid sequences with other eukaryotic species. The recombinant CcCK2α and CcCK2β proteins were purified by affinity chromatography to homogeneity, after overexpression in Escherichia coli. CcCK2α is capable to utilize GTP and its activity and is inhibited by polyanions and stimulated by polycations in phosphorylation assays, using purified acidic ribosomal protein P1 as a substrate.
authors: Department of Genetics and Biotechnology, University of Athens, Panepistimiopolis, Athens 15701, Greece. skougian@biol.uoa.gr
authors: Sophia
authors: S
authors: Kouyanou-Koutsoukou
authors: Andrea
authors: A
authors: Baier
authors: Regina-Maria
authors: RM
authors: Kolaitis
authors: Evanthia
authors: E
authors: Maniatopoulou
authors: Konstantina
authors: K
authors: Thanopoulou
authors: Ryszard
authors: R
authors: Szyszka
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D047390:Casein Kinase II / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D020134:Catalytic Domain; D033622:Ceratitis capitata / Q000201:enzymology* / Q000235:genetics*; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D055767:Immobilized Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification*; D016415:Sequence Alignment
chemical_list: D018076:DNA, Complementary; D055767:Immobilized Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D047390:Casein Kinase II