OCT-1 POU DOMAIN DNA INTERACTIONS - COOPERATIVE BINDING OF ISOLATED SUBDOMAINS AND EFFECTS OF COVALENT LINKAGE

Structural and biochemical studies of Oct-1 POU domain-DNA interaction s have raised important questions about cooperativity and the role of the linker connecting the POU-specific domain and the POU homeo domain . To analyze these interactions, we have studied binding of the isolat ed domains. Surprisingly, we find that two unlinked polypeptides corre sponding to the POU-specific domain and the POU homeo domain bind coop eratively to the octamer site and have a coupling energy of 1.6 kcal/m ole. We suggest that overlapping DNA contacts near the center of the o ctamer site may be the source of this cooperativity, as there are no p rotein-protein contacts between the domains in the crystal structure o f the Oct-1 POU domain-DNA complex. These studies also have allowed us to describe the thermodynamic contribution of the linker (present in the intact POU domain) in terms of an effective concentration (3.6 mM) . The broader implications for understanding cooperativity in protein- DNA recognition and gene regulation are discussed.