Characterization of a purified chromatin acceptor protein (receptor binding factor 1) for the avian oviduct progesterone receptor.
The specific, high-affinity binding of the avian oviduct progesterone receptor (PR) with target-cell nuclei and chromatin has been shown to involve DNA complexed with specific chromatin acceptor proteins. One of these chromatin acceptor proteins has been partially purified and found to be a small hydrophobic protein with a broad pI of 5.0-6.0 [Goldberger, A., & Spelsberg, T. C., (1988) Biochemistry 27, 2103-2109]. This paper describes the final purification over 100,000-fold to apparent homogeneity of this candidate PR acceptor protein, termed the receptor binding factor 1 (RBF-1). When the avian genomic DNA is bound by RBF-1, saturable, high-affinity (KD approximately 2 x 10(-9) M) binding sites for PR are generated. RBF-1 has a unique, hydrophobic N-terminal sequence. The PR binding to the RBF-1-DNA complexes is shown to be dependent on an intact activated PR with which excess nonradiolabeled PR can compete. By use of a new, highly specific monoclonal antibody (mAb) to the RBF-1 with Western immunoblotting, RBF-1 was shown to be localized in the nucleus and to be tissue and species specific. Selective removal of the chromatin proteins containing RBF-1 results in the loss of the highest affinity class of PR binding sites. A second class of residual PR binding sites remains in the nucleoacidic protein (NAP), a complex of proteins more tightly bound to the DNA. This class of PR binding activity has been classified as the RBF-2. The RBF-1 is estimated to be 0.03% of the total chromatin protein with about 1.2 x 10(5) molecules/diploid cell.
Schuchard
M
M
Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota 55905.
Rejman
J J
JJ
McCormick
D J
DJ
Gosse
B
B
Ruesink
T
T
Spelsberg
T C
TC
D000595:Amino Acid Sequence; D000818:Animals; D030161:Avian Proteins; D001667:Binding, Competitive; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D002645:Chickens; D002843:Chromatin / Q000737:chemistry*; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D010057:Oviducts / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*
D030161:Avian Proteins; D002352:Carrier Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; C085646:RBF-1 protein, Gallus gallus; D011980:Receptors, Progesterone; D004247:DNA