Bailey S, Wichitwechkarn J, Johnson D, Reilly B E, Anderson D L, Bodley J W
Department of Biochemistry, University of Minnesota, Minneapolis 55455.
J Biol Chem. 1990 Dec 25;265(36):22365-70.
An unusual RNA molecule encoded by the Bacillus subtilis bacteriophage phi 29 is a structural component of the viral prohead and is required for the ATP-dependent packaging of DNA. Here we report a model of secondary structure for this prohead RNA developed from a phylogenetic analysis of the primary sequences of prohead RNAs of related phages. Twenty-nine phages related to phi 29 were found to produce prohead RNAs. These RNAs were analyzed by their ability to replace phi 29 RNA in in vitro phage assembly, by Northern blot hybridization with a probe complementary to phi 29 RNA, and by partial and complete sequence analyses. These analyses revealed four quite different sequences ranging in length from 161 to 174 residues. The secondary structure deduced from these sequences, in agreement with earlier observations, indicated that prohead RNA is organized into two domains. The larger 5'-domain (Domain I) is composed of 113-117 residues and contains four helices. Three of these helices appear to be organized into a central stem that is interrupted by two unpaired loops and the fourth helix and loop. The smaller 3'-domain (Domain II) is composed of 40-44 residues and consists of two helices. Domains I and II are separated by 8-13 unpaired residues. Nuclease cleavage occurs readily in this single-stranded joining region, and this cleavage allows the subsequent separation of the two RNA domains. The separated Domain I is fully active in DNA packaging in vitro. The functional significance and biological role of Domain II are unknown. The phylogenetic secondary structure model provides a basis for further analysis of the role of this RNA in bacteriophage morphogenesis.
由枯草芽孢杆菌噬菌体 phi 29 编码的一种异常 RNA 分子是病毒原头部的结构成分，是 DNA 依赖 ATP 进行包装所必需的。在此，我们报告了一种基于对相关噬菌体原头部 RNA 一级序列的系统发育分析而构建的该原头部 RNA 二级结构模型。发现 29 种与 phi 29 相关的噬菌体可产生原头部 RNA。通过它们在体外噬菌体组装中替代 phi 29 RNA 的能力、与 phi 29 RNA 互补探针的 Northern 印迹杂交以及部分和完整序列分析，对这些 RNA 进行了分析。这些分析揭示了四种长度在 161 至 174 个残基之间的截然不同的序列。从这些序列推导的二级结构与早期观察结果一致，表明原头部 RNA 被组织成两个结构域。较大的 5' 结构域（结构域 I）由 113 - 117 个残基组成，包含四个螺旋。其中三个螺旋似乎被组织成一个中央茎，该中央茎被两个未配对的环以及第四个螺旋和环打断。较小的 3' 结构域（结构域 II）由 40 - 44 个残基组成，由两个螺旋组成。结构域 I 和 II 由 8 - 13 个未配对的残基分隔。核酸酶在这个单链连接区域容易发生切割，这种切割使得随后两个 RNA 结构域能够分离。分离后的结构域 I 在体外 DNA 包装中具有完全活性。结构域 II 的功能意义和生物学作用尚不清楚。系统发育二级结构模型为进一步分析这种 RNA 在噬菌体形态发生中的作用提供了基础。
