Identification of nonhistone chromosomal proteins binding specifically to cloned alpha-fetoprotein DNA sequences

A sequential nitrocellulose filter binding assay has been used to quantitate specific binding by nonhistone proteins of a cloned DNA fragment coding for the 3' end of the mouse alpha-fetoprotein gene. Nonhistone proteins from fetal mouse liver, from adult liver, and from kidney, all bind this fragment specifically; there is no significant difference in binding by proteins isolated from these three tissues. The proteins responsible for specific binding have been isolated by sucrose density gradient ultracentrifugation of DNA-protein complexes and characterized by sodium dodecyl sulfate - polyacrylamide gel electrophoresis and electron microscopy. The majority of the specific binding proteins from adult liver consists of two polypeptides of molecular weights 53 500 and 51 000; three or four minor bands in the molecular weight range 43 000-48 000 are also observed. Similar specific binding proteins exist among the fetal liver nonhistone proteins. The native binding proteins as visualized by electron microscopy are very large and appear to bind specifically to a site near the centre of the alpha-fetoprotein DNA fragment, as well as nonspecifically to the ends of all DNA molecules.