X-ray Crystal Structural Analysis of Bacterial Iron Binding Protein and Development for New Antibiotics

点击翻译按钮获取中文摘要

In this study, we investigated the X-ray structural analyses on the iron-binding proteins(Dps (DNA protection during starvation) protein and ferritin protein) from peridontopathic bacteria, Actinobacillus actinomycetemcomitans, which play an important role in protecting cellular macromolecules from damage by reactive oxygen species.A.actinomycetemcomitans expressed two ferritin protein(Ftn1 and Ftn2). The Ftn 1 could be successfully crystallized, but Ftn 2 became only amorphous form. Crystals of the Dps-like protein of A. actinomycetemcomitans were grown by the hanging drop method of vapour diffusion from a solution containing 28% polyethylene glycol 400 in 0.1 M HEPES-Na buffer pH 7.5 with 0.2 M calcium chloride dihydrate. Small but perfect hexagonal rods were grown with a protein concentration of~10 mg/ml. These rods diffract strongly to 1.9 A in-house and were found to be in the hexagonal space group P63 with cell dimensions a = b = 128.5 A, c = 91.lA and γ=120°. A 93.9% complete da … More ta set was collected to 1.9 A with a multiplicity of 2.9 and an R_<merge> of 0.071. The structure of the Dps-like protein of A.actinomycetemcomitans was solved by molecular replacement using the dodecameric ferritin like protein of Listeria innocua as the search model. The asymmetric unit contains four unique subunits arranged around the crystallographic 3-fold axis.We also analyzed the antimicrobial activity against oral bacteria of human beta-defensin-2 (hBD-2) as a candidate of new antimicrobial substances. hBD-2 shouwed antimicrobial activity gainst Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Streptococcus mutans, which was approximately equal to that of minocycline at equimolar concentrations. And we further examined the effect of ingredients and urinary metabolites of cranberry, which is rich in polyphenols, has been found to have various effects beneficial to human health, on biofilm formation by Escherichia coli. We found that ferulic acid, homovanillic acid, 4-coumaric acid, isoferulic acid and vanillic acid with inhibitory activity on Escherichia coli biofilm formation. Less