Tomohide Uno, Yuka Nomura, Masahiko Nakamura, Atsushi Nakao, Shoji Tajima, Kengo Kanamaru, Hiroshi Yamagata, Yousuke Iwanaga
Journal of Insect Science 5 (8), 1-8, (1 March 2005) https://doi.org/10.1673/031.005.0801
KEYWORDS: DNA methylation, phosphorylation
A cDNA clone encoding methyl DNA binding domain-containing protein (bMBD2/3) was obtained by homology searches using a Bombyx mori fat body cDNA library. The cDNA encoded a polypeptide with 249 amino acids sharing 54% similarity with the methyl DNA binding protein from Drosophila melanogaster. To characterize the biochemical properties of bMBD2/3, the clone was expressed in Escherichia coli as His-tagged protein. The recombinant protein was purified to homogeneity using Ni-NTA superflow resin and heparin agarose. The protein showed specific methyl DNA binding activity and was phosphorylated by protein kinase in vitro. Immunoblotting using the purified antibody indicated that bMBD2/3 was expressed in almost all tissues. Using west-western blotting analysis, some proteins that interact with bMBD2/3 were identified in the brain. This is the first report that insect MBD is phosphorylated and is present in adult tissues. These results suggest that bMBD2/3 plays important roles in the DNA methylation-specific transcription of Bombyx mori.