In-vitro phosphorylation activity by recombinant alpha and beta subunits of Bombyx mori casein kinase 2.
To clarify the control mechanism of the catalytic activity of casein kinase 2 (CK2) during early embryonic development in the silkworm, Bombyx mori, we attempted an in-vitro functional analysis by using the recombinant alpha and beta subunits of B. mori CK2 (rBmCK2alpha and rBmCK2beta) produced in a bacterial system. The renatured rBmCK2alpha possessed protein kinase activity. When rBmCK2alpha and rBmCK2beta were reconstituted in an approximate 1:1 molar ratio, the catalytic activity was almost the same as that of rBmCK2alpha alone. The catalytic activity of rBmCK2alpha was inhibited by polylysine, which is one of the activators of CK2 activity. However, when using the reconstituted rBmCK2alpha and rBmCK2beta (rBmCK2), activation by polylysine was observed. We examined the influence of sorbitol and 3-hydroxykynurenine (3-OHK), which are contained mainly in diapause eggs, on the phosphorylation activity of rBmCK2. Three-OHK inhibited rBmCK2 activity, but sorbitol had no effect on it. Furthermore, a functional analysis using rBmCK2alpha and beta subunits of Drosophila melanogaster CK2 revealed that a difference in the C-terminal amino acid of the CK2beta subunit influenced the phosphorylation activity of rBmCK2alpha. These results may provide new insights for clarifying the control mechanism of B. mori casein kinase 2 in eggs.
Yamamoto
Takayuki
T
Biological Laboratory, College Of Liberal Arts and Sciences, Kitasato University, Kitasato, Sagamihara, Japan.
Sawada
Hiroshi
H
D000595:Amino Acid Sequence; D000818:Animals; D012831:Bombyx / Q000196:embryology / Q000201:enzymology* / Q000235:genetics; D047390:Casein Kinase II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D010063:Ovum / Q000201:enzymology* / Q000254:growth & development; D010766:Phosphorylation; D010770:Phosphotransferases; D011494:Protein Kinases / Q000378:metabolism; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D029721:Drosophila Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D010770:Phosphotransferases; D011494:Protein Kinases; D047390:Casein Kinase II