Characterization of the binding of spike H protein of bacteriophage φX74 with receptor lipopolysaccharides
Characterization of the binding of spike H protein of bacteriophage φX74 with receptor lipopolysaccharides
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DOI：
10.1093/oxfordjournals.jbchem.a022643
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发表时间：
2000-04-01
影响因子：
2.7
通讯作者：
Kashimura, N
中科院分区：
文献类型：
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作者：
Inagaki, M;Tanaka, A;Kashimura, N
将噬菌体phi X174的刺突H蛋白制备为六组氨酸标记的融合体（HisH）。在酶联板分析中，HisH与phi X174敏感菌株（大肠杆菌C和鼠伤寒沙门氏菌Ra）的脂多糖（LPS）特异性结合，而与phi X174不敏感菌株（大肠杆菌C和鼠伤寒沙门氏菌Ra）的LPS结合较弱。coli F583（Rd（2））和E. coliO 111：B4（光滑株），其R-核心上有额外的O-重复序列。用荧光滴定法测定了HisH与LPS的结合平衡，测定了解离常数Kd为7.02 ± 0.37 μ M，吉布斯自由能变化Δ G（0）为-29.1kJmol（-1）。（在22 ° C，pH 7.4）。基于范特霍夫图中的Kd的温度依赖性，计算出标准焓变Δ H-0和熵变Δ S-0在22 ° C分别为+23.7 kJ mol（-1）和179 J mol（-1）K-1，由此推断这种结合是熵驱动的反应。
The spike H protein of bacteriophage phi X174 was prepared as a hexa histidine-tagged fusion (HisH). On enzyme-linked plate assaying, HisH was found to bind specifically to the lipopolysaccharides (LPSs) of phi X174-sensitive strains, Escherichia coli C and Salmonella typhimurium Ra chemotype, having the complete oligosaccharide sequence of the R-core on the LPSs, In sharp contrast, HisH bound weakly to the LPSs of phi X174-insensitive strains, i.e. E. coli F583 (Rd(2)) lacking some terminal saccharides and E. coli O111:B4 (smooth strain) having additional O-repeats on the R-core, The fluorescence spectra of HisH changed dose-dependently in the case of the LPS of E. coli C, the intensity increasing and the emission peak shifting to the shorter wavelength side, which was attributable to the hydrophobic interaction of HisH with the LPS, The binding equilibrium was analyzed by fluorometric titration to determine the dissociation constant K-d, 7.02 +/- 0.37 mu M, and the Gibbs free energy change Delta G(0), -29.1 kJ mol(-1) (at 22 degrees C, pH 7.4), Based on the temperature dependence of K-d in a van't Hoff plot, the standard enthalpy change Delta H-0 and the entropy change Delta S-0 were calculated to be +23.7 kJ mol(-1) and 179 J mol(-1) K-1 at 22 degrees C, respectively, and this binding was thereby concluded to be an entropy-driven reaction.