SOLUTION STRUCTURE OF A POU-SPECIFIC HOMEODOMAIN - 3D-NMR STUDIES OF HUMAN B-CELL TRANSCRIPTION FACTOR OCT-2

The POU DNA-binding motif defines a conserved family of eukaryotic tra nscription factors involved in regulation of gene expression. This bip artite motif consists of an N-terminal POU-specific domain (POUs), a f lexible linker, and a C-terminal POU-specific homeodomain (POUHD) Here we describe the solution structure of a POU-specific homeodomain. An NMR model is obtained from Oct-2, a human B-cell specific transcriptio n factor which participates in the regulation of immunoglobulin genes. A fragment of Oct-2 containing POUHD and an adjoining linker was expr essed in Escherichia coli and characterized by three-dimensional nucle ar magnetic resonance (3D-NMR) spectroscopy. Complete H-1 and N-15 res onance assignment of the POUHD moiety is presented. The POUHD solution structure, as calculated by distance geometry and simulated annealing (DG/SA), is similar to that of canonical homeodomains. A salient diff erence between solution and crystal structures is observed in the C-te rminal segment of alpha-helix 3 (the HTH recognition helix), which is not well ordered in solution. Because this segment presumably folds up on specific DNA binding, its flexibility in solution may reduce the in trinsic DNA affinity of POUHD in the absence of POUs.