Kralicek A V, Vesper N A, Ralston G B, Wake R G, King G F
Department of Biochemistry, University of Sydney, NSW, Australia.
Biochemistry. 1993 Sep 28;32(38):10216-23. doi: 10.1021/bi00089a043.
We have used analytical ultracentrifugation in combination with a number of spectroscopic techniques to analyze the symmetry and secondary structure of the DNA-binding replication terminator protein (RTP) of Bacillus subtilis. Sedimentation equilibrium studies confirm that RTP is a dimer in solution under the conditions used for spectroscopic analysis, whereas the number of cross peaks displayed in 1H-15N HSQC NMR spectra of uniformly 15N-labeled RTP are consistent with the primary structure of the monomer. These two results in combination lead to the conclusion that RTP is a symmetric dimer in solution. Circular dichroic and Fourier-transform infrared spectra reveal, in contrast to the results obtained from a number of commonly used secondary structure prediction algorithms, that RTP contains 20-30% alpha-helical and 40-50% beta-sheet/beta-turn secondary structure and that the conformation of the protein remains unchanged over the pH range 5-8. It is proposed on the basis of protein folding-class prediction algorithms, in combination with various physical properties of RTP, that it belongs to the alpha + beta protein-folding class.
我们运用分析超速离心技术，并结合多种光谱技术，来分析枯草芽孢杆菌DNA结合复制终止蛋白（RTP）的对称性和二级结构。沉降平衡研究证实，在用于光谱分析的条件下，RTP在溶液中是二聚体，而均匀15N标记的RTP的1H-15N HSQC NMR谱中显示的交叉峰数量与单体的一级结构一致。这两个结果共同得出结论，即RTP在溶液中是对称二聚体。圆二色光谱和傅里叶变换红外光谱显示，与许多常用二级结构预测算法得出的结果相反，RTP含有20%-30%的α-螺旋和40%-50%的β-折叠/β-转角二级结构，并且该蛋白质的构象在pH值5-8范围内保持不变。基于蛋白质折叠类预测算法，并结合RTP的各种物理性质，推测它属于α+β蛋白质折叠类。
