Structure of an antigen-binding fragment bound to stem-loop DNA and crystallization of recombinant haemophilus influenzae e(P4) acid phosphatase
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DNA-1 and 11F8 are anti-ss DNA antibodies derived from autoimmune lupus-prone mice. They are very similar to each other in terms of CDR sequence and preference for binding T-rich ss DNA. G1-17 is an oligonucleotide identified by in vitro selection experiments and binds with high affinity and specificity to Fab 11F8. G5-14 is a synthetic oligonucleotide with the tennucleotide sequence identical to the stem-loop portion above the bulge of G1-17. The 1.95 A ̊resolution DNA-1/G5-14 structure shows that the two DNA strands dimerize to form a double stranded DNA dumbbell and have a large conformational change including the breaking and reformation of hydrogen bonds. The most striking feature of the Fab/DNA interactions is the use of extensive [pi-pi] stacking of the DNA bases and the protein side chains. These results provide insights into the specific recognition model of anti-DNA Abs and the potential challenges in structure based drug design to treat autoimmune diseases. The second part of this thesis describes purification and crystallization of Haemophilus influenzae class C acid phosphatise P4, and acquisition of a 1.7 Å ̊resolution native X-ray diffraction data set. The space group of the crystals is P4₂2₁2 with a = 65.6, c = 101.4 A ̊one protein molecule per asymmetric unit and 37 % solvent content. This is the first report of crystallization of a class C acid phosphatase.