Purification and characterization of W-protein. A DNA-binding protein showing high affinity for the W chromosome-specific repetitive DNA sequences of chicken.
A protein component, which binds with high affinity to the W chromosome-specific XhoI family repetitive DNA of chicken (Tone, M., Sakaki, Y., Hashiguchi, T., and Mizuno, S. (1984) Chromosoma (Berl.) 89, 228-237), was detected in the 0.35 M NaCl extract of the female chicken liver nuclei. This protein, designated as W-protein, was substantially purified by phosphocellulose, hydroxyapatite, and DEAE-Toyopearl column chromatography. Molecular weight of W-protein was estimated to be about 72,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but it seems to form multimeric structure having apparent molecular weight of about 2.3 X 10(6) under nondenaturing conditions. W-Protein binds strongly to both 0.7- and 1.1-kb repeating units of the XhoI family, both of which show curved DNA characteristics, and weakly to the AATAT-satellite sequence of Drosophila melanogaster. Stable binding of W-protein requires greater than or equal to 300 base pairs of the 0.7-kilobases sequence, or more than 14 tandem repeats of the 21-base pair internal repeating unit of the 0.7-kilobase sequence. DNA footprint analysis and effects of some DNA-binding compounds suggest that the DNA double helix wraps around W-protein or its multimeric form contacting through A-T-rich minor grooves. A possible role of W-protein in the formation of W heterochromatic body is discussed.
Harata
M
M
Department of Agricultural Chemistry, Tohoku University, Sendai, Japan.
Ouchi
K
K
Ohata
S
S
Kikuchi
A
A
Mizuno
S
S
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000032:analysis; D002645:Chickens; D002845:Chromatography; D002875:Chromosomes / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D004262:DNA Restriction Enzymes; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins; D003850:Deoxyribonuclease I / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D008099:Liver / Q000032:analysis*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012091:Repetitive Sequences, Nucleic Acid
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D004247:DNA; D004262:DNA Restriction Enzymes; D003850:Deoxyribonuclease I