Crystal structure of paired domain-DNA complex

This thesis describes the determination of a paired domainDNA complex crystal structure (involving the paired domain of the Drosophila Prd protein), and discusses the structural basis of DNA binding specificity of the paired domain and the structural basis of Pax developmental mutations. It also describes the cocrystallization of the human PAX6 paired domain-DNA complex. Chapter 1 provides an introduction to paired domains and the Pax family. Pax genes play very important roles for vertebrate development. Mutations in several Pax genes have been associated with mouse and human congenital disorders. The paired domain, a highly conserved DNA-binding domain, is critical for Pax protein function. Chapter 2 describes the purification of Drosophila Prd paired domain, the crystallization of the Prd paired domain-DNA complex, and the determination of the crystal structure of this complex. Chapter 3 describes the structure of the Prd paired domain DNA complex. The crystal structure shows that the paired domain folds as two independent sub-domains, each containing a helical structure that is very similar to the homeodomain. The N-terminal domain makes extensive DNA contacts. It has a novel -turn motif that fits in the minor groove and a HTH unit that contacts the major groove. The -turn makes base specific contacts in the minor groove, and is critical for both DNA binding and for Pax in vivo function. The HTH unit folds like a homeodomain but docks on DNA like repressor. The C-terminal domain of the Prd paired domain does not contact the optimized DNA binding site, and other experiments have shown that it is not required for DNA recognition.