Production of HMG-3 by limited trypsin digestion of purified high-mobility-group nonhistone chromatin proteins.
Three isolated nonhistone proteins (HMG-1, HMG-2 and HMG-E) have been purified from chicken erythrocyte chromatin without exposure to overt denaturing conditions, and subjected to limited proteolysis. When treated with trypsin, the three proteins exhibited similar patterns of degradation, as judged by SDS and acid/urea gel electrophoresis. In particular, the first product, P1 (a relatively stable intermediate in each digestion), was a protein analogous to HMG-3, a principal degradation product in preparations of calf thymus high-mobility-group proteins. At least in the case of HMG-E, the products formed by tryptic attack on P1 are the two individual DNA binding domains of HMG-E. P1 derived from HMG-E and one of the individual DNA binding domains of HMG-E were purified by chromatography on columns containing DNA-cellulose or phosphocellulose. The properties of these two portions of HMG-E are consistent with our recently postulated three-domain structure for HMG-1 and its homologs (Reeck, G.R., Isackson, P.J. and Teller, D.C. (1982) Nature 300, 76-78). Thus, P1 consists of two DNA-binding domains of approximately equal molecular weight covalently linked together. From chromatography on DNA-cellulose columns, it is clear that P1 binds to DNA more tightly than does HMG-E. The highly acidic C-terminal domain of HMG-E (which is removed by trypsin in generating P1) thus counteracts the DNA binding of the two other domains of HMG-E (at least in the protein's interaction with purified DNA).
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002645:Chickens; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006609:High Mobility Group Proteins; D008970:Molecular Weight; D014357:Trypsin / Q000378:metabolism*
D000596:Amino Acids; D002868:Chromosomal Proteins, Non-Histone; D006609:High Mobility Group Proteins; D004247:DNA; D014357:Trypsin