The barrier-to-autointegration factor BAF binds to the LEM domain (Em LEM ) of the nuclear envelope protein emerin and plays an essential role in the nuclear architecture of metazoan cells. In addition, the BAF 2 dimer bridges and compacts doublestranded DNA nonspecifically via two symmetry-related DNA binding sites. In this article we present biophysical and structural studies on a complex of BAF 2 and Em LEM . Light scattering, analytical ultracentrifugation, and NMR indicate a stoichiometry

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... f one molecule of Em LEM bound per BAF 2 dimer. The equilibrium dissociation constant (K d ) for the interaction of the BAF 2 dimer and Em LEM , determined by isothermal titration calorimetry, is 0.59 ؎ 0.03 M. Z-exchange spectroscopy between corresponding cross-peaks of the magnetically non-equivalent subunits of the BAF 2 dimer in the complex yields a dissociation rate constant of 78 ؎ 2 s ؊1 . The solution NMR structure of the BAF 2 -Em LEM complex reveals that the LEM and DNA binding sites on BAF 2 are non-overlapping and that both subunits of the BAF 2 dimer contribute approximately equally to the Em LEM binding site. The relevance of the implications of the structural and biophysical data on the complex in the context of the interaction between the BAF 2 dimer and Em LEM at the nuclear envelope is discussed. The barrier to autointegration factor (BAF) 2 (1) and the inner nuclear envelope LEM-domain protein emerin (2) are highly conserved cellular proteins throughout the metazoan kingdom that play an important role in nuclear architecture (3). BAF is an all-helical obligate dimer (4) that possesses two symmetry related DNA binding sites that permit BAF to bridge DNA chains and thereby compact DNA (5). Emerin is a member of