11] DNA-binding proteins Sac7d and Sso7d from Sulfolobus

Abstract

The chromatin of the archaea Sulfolobus acidocaldarius and Sulfolobus solfataricus contains a number of small basic proteins ranging in molecular weight from 7000 to 10,000. The 7-kDa proteins have been the most studied and believed to be important in DNA compaction and stabilization of duplex DNA at the growth temperatures of 75° for S. acidocaldarius and 80° for S. solfataricus. The 7-kDa chromatin proteins of S. acidocaldarius consist of five species, designated Sac7a, b, c, d, and e in order of increasing basicity. Sac7d (molecular weight 7477) and Sac7e (molecular weight 7338) differ by six amino acid residues and are coded by distinct genes. Sac7a and b are carboxy-terminal truncated forms of the Sac7d. Only one form of Sso7 has been characterized; it is referred to as Sso7d (MW 7019) based on homology to Sac7d. The structures of both Sac7d and Sso7d DNA-protein complexes have also been determined by X-ray Crystallography. This chapter describes the purification of native Sac7 and Sso7 from Sulfolobus and recombinant proteins from E. coli, along with a brief description of various physical properties of the purified proteins. Assays for DNA binding and RNase activities are described and evaluated.