THE CRYSTAL-STRUCTURE OF A HYPERTHERMOPHILIC ARCHAEAL TATA-BOX BINDING-PROTEIN

This study analyzes the three-dimensional structure of the TATA-box bi nding protein (TBP) from the hyperthermophilic archaea Pyrococcus woes ei. The crystal structure of P. woesei TBP (PwTBP) was solved at 2.2 A ngstrom by X-ray diffraction and as expected from sequence homology (3 6% to 41% identical to eukaryotic TBPs) its overall structure is very similar to eukaryotic TBPs. The thermal unfolding transition temperatu re of this protein was measured by differential scanning calorimetry t o be 101 degrees C, which is more than 40 degrees C higher than that o f yeast TBP. Preliminary titration calorimetry data show that the affi nity of PwTBP for its DNA target, unlike its eukaryotic counterparts, is enhanced by increasing the temperature and salt concentration. The structure reveals possible explanations for this thermostability and t hese unusual DNA binding properties. The crystal structure of this hyp erthermostable protein was compared to its mesophilic homologs and ana lyzed for differences in the native structure that may contribute to t hermostability. Differences found were: (1) a disulfide bond not found in mesophilic counterparts; (2) an increased number of surface electr ostatic interactions; (3) more compact protein packing. The presumed D NA binding surface of PwTBP, like its eukaryotic counterparts, is hydr ophobic but the electrostatic profile surrounding the protein is relat ively neutral compared to the asymmetric positive potential that surro unds eukaryotic TBPs. The total reliance on a hydrophobic interface wi th DNA may explain the enhanced affinity of PwTBP for its DNA promoter at higher temperatures and increased salt concentration. (C) 1996 Aca demic Press Limited