Williamson D L, Renaudin J, Bové J M
Department of Anatomical Sciences, State University of New York, Stony Brook 11794-8081.
J Bacteriol. 1991 Jul;173(14):4353-62. doi: 10.1128/jb.173.14.4353-4362.1991.
Electron microscopic observation of spiroplasmas lysed by detergent (sodium deoxycholate) revealed the release of bundles of fibrils from the cells. Individual fibrils are 4 nm in diameter and possess a 9-nm periodicity along their length. These fibrils are thought to function as cytoskeletal structures involved in the shape and motility of spiroplasmas. Polyacrylamide gel electrophoresis of density gradient-purified fibrils showed a protein of approximately 55 kDa. Oligonucleotide probes were constructed from the N-terminal amino acid sequence of two peptides obtained after V8 protease hydrolysis of the fibril protein. The probes were used to identify the clones in a genomic DNA library of Spiroplasma citri that contained inserts carrying the probe sequence. Sequencing of a 3.3-kbp fragment yielded the full open reading frame of the fibril protein gene and the start of a second open reading frame of an unknown protein. The fibril protein is composed of 515 amino acids, which have a computed molecular mass of 59 kDa. Northern (RNA) blot hybridization and primer extension experiments showed that transcription of the fibril protein gene starts from a promoter located 100 nucleotides upstream of the initiation codon and stops at a rho-independent type terminator, leading to a 1.7-kbp transcript. Southern blot hybridization of genomic DNA using the fibril protein gene as the probe showed that a single copy of the gene is present in the chromosomes of both S. citri and Spiroplasma melliferum. The genotypic symbol fib is proposed for the spiroplasma fibril protein gene.
对被去污剂（脱氧胆酸钠）裂解的螺原体进行电子显微镜观察，发现细胞释放出成束的纤丝。单个纤丝直径为4纳米，沿其长度具有9纳米的周期性。这些纤丝被认为起着细胞骨架结构的作用，参与螺原体的形态和运动。对密度梯度纯化的纤丝进行聚丙烯酰胺凝胶电泳，显示出一种约55千道尔顿的蛋白质。根据纤丝蛋白经V8蛋白酶水解后获得的两种肽的N端氨基酸序列构建了寡核苷酸探针。这些探针用于鉴定柑桔螺原体基因组DNA文库中含有携带探针序列插入片段的克隆。对一个3.3千碱基对片段进行测序，得到了纤丝蛋白基因的完整开放阅读框以及一个未知蛋白的第二个开放阅读框的起始部分。纤丝蛋白由515个氨基酸组成，计算分子量为59千道尔顿。Northern（RNA）印迹杂交和引物延伸实验表明，纤丝蛋白基因的转录起始于起始密码子上游100个核苷酸处的一个启动子，并在一个不依赖ρ因子的终止子处停止，产生一个1.7千碱基对的转录本。用纤丝蛋白基因作为探针进行基因组DNA的Southern印迹杂交表明，该基因在柑桔螺原体和蜜蜂螺原体的染色体中均以单拷贝存在。建议将螺原体纤丝蛋白基因的基因型符号定为fib。
