Purification and properties of yolk protein factor I, a sequence-specific DNA-binding protein from Drosophila melanogaster.
We report the purification and some of the biochemical properties of yolk protein factor I (YPF1). This protein binds to a specific site in the yolk protein 1 gene (yp1) of Drosophila melanogaster. YPF1 has been purified to 95% homogeneity and consists of a heterodimer of two subunits with molecular weights 85,000 and 69,000. The protein is highly asymmetric with a frictional ratio of 1.56 which leads to calculated dimensions of 510 x 51 A when modeled as a prolate ellipsoid of revolution. It binds the yp1 DNA site with a protein/DNA stoichiometry of 1:1. Binding to that site is essentially irreversible with a dissociation rate constant of koff less than or equal to 2 x 10(-7) s-1, which gives the complex a dissociation half-life of approximately 55 days. The measured apparent second order association rate constant is 4 x 10(8) M-1 s-1 resulting in a calculated equilibrium dissociation constant of KD less than or equal to 5 x 10(-16) M. YPF1 also has a 10(8) selectivity for the yp1 site over poly(dA).poly(dT) (KDapp = 2 x 10(-8) M(nucleotide].
Mitsis
P G
PG
Rosenstiel Center, Brandeis University, Waltham, Massachusetts 02254-9110.
Wensink
P C
PC
D000818:Animals; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000032:analysis* / Q000235:genetics; D004527:Egg Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D011487:Protein Conformation; D012045:Regulatory Sequences, Nucleic Acid
D004268:DNA-Binding Proteins; D004527:Egg Proteins; D004247:DNA