Nakamura S, Ohta S, Arai K, Arai N, Oshima T, Kaziro Y
Eur J Biochem. 1978 Dec;92(2):533-43. doi: 10.1111/j.1432-1033.1978.tb12775.x.
Molecular properties of the polypeptide chain elongation factors from Thermus thermophilus HB8 have been investigated and compared with those from Escherichia coli. 1. As expected, the factors purified from T. thermophilus were exceedingly heat-stable. Even free EF-Tu not complexed with GDP was stable after heating for 5 min at 60 degrees C. 2. GDP binding activity of T. thermophilus EF-Tu was also stable in various protein denaturants, such as 5.5 M urea, 1.5 M guanidine-HCl, and 4 M LiCl. 3. Amino acid compositions of EF-Tu and EF-G from T. thermophilus were similar to those from E. coli. On the other hand, amino acid composition of T. thermophilus EF-Ts was considerably different from that of E. coli EF-Ts. 4. In contrast to E. coli EF-Tu, T. thermophilus EF-Tu contained no free sulfhydryl group, but one disulfide bond. The disulfide bond was cleaved by sodium borohydride or sodium sulfite under native conditions. The heat stability of the reduced EF-Tu . GDP, as measured by GDP binding activity, did not differ from that of the untreated EF-Tu . GDP. 5. T. thermophilus EF-Ts contained, in addition to one disulfide bond, a sulfhydryl group which could be titrated only after complete denaturation of the protein. 6. Under native conditions one sulfhydryl group of T. thermophilus EF-G was titrated with p-chloromercuribenzoate, while the rate of reaction was very sluggish. The sulfhydryl group appears to be essential for interaction with ribosomes, whereas the ability to form a binary GDP . EF-G complex was not affected by its modification. The protein contained also one disulfide bond. 7. Circular dichroic spectra of EF-Tu from T. thermophilus and E. coli were very similar. Binding of GDP or GTP caused a similar spectral change in both. T. thermophilus and E. coli EF-Tu. On the other hand, the spectra of T. thermophilus EF-G and E. coli EF-G were significantly different, the content of ordered structure being higher in the former as compared to the latter.
嗜热栖热菌HB8多肽链延伸因子的分子特性已被研究，并与大肠杆菌的进行了比较。1. 正如预期的那样，从嗜热栖热菌中纯化的因子具有极高的热稳定性。即使是未与GDP结合的游离EF-Tu，在60℃加热5分钟后仍保持稳定。2. 嗜热栖热菌EF-Tu的GDP结合活性在各种蛋白质变性剂中也很稳定，如5.5M尿素、1.5M盐酸胍和4M LiCl。3. 嗜热栖热菌的EF-Tu和EF-G的氨基酸组成与大肠杆菌的相似。另一方面，嗜热栖热菌的EF-Ts的氨基酸组成与大肠杆菌的EF-Ts有很大不同。4. 与大肠杆菌EF-Tu不同，嗜热栖热菌EF-Tu不含游离巯基，但含有一个二硫键。在天然条件下，二硫键可被硼氢化钠或亚硫酸钠裂解。通过GDP结合活性测定，还原后的EF-Tu·GDP的热稳定性与未处理的EF-Tu·GDP没有差异。5. 嗜热栖热菌EF-Ts除含有一个二硫键外，还含有一个巯基，该巯基只有在蛋白质完全变性后才能被滴定。6. 在天然条件下，嗜热栖热菌EF-G的一个巯基可用对氯汞苯甲酸滴定，但其反应速率非常缓慢。该巯基似乎对与核糖体的相互作用至关重要，而形成二元GDP·EF-G复合物的能力不受其修饰的影响。该蛋白质还含有一个二硫键。7. 嗜热栖热菌和大肠杆菌的EF-Tu的圆二色光谱非常相似。GDP或GTP的结合在两者中引起相似的光谱变化。嗜热栖热菌和大肠杆菌的EF-Tu。另一方面，嗜热栖热菌EF-G和大肠杆菌EF-G的光谱有显著差异，前者的有序结构含量比后者高。
