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Molecular characterization and protein analysis of the cap region, which is essential for encapsulation in Bacillus anthracis.

机译:帽状区域的分子表征和蛋白质分析,这对于封装在炭疽芽胞杆菌中至关重要。

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摘要

By using genetic complementation tests with various in vitro-constructed mutants with mutations in the cap region (which is essential for encapsulation in Bacillus anthracis), we identified three cistrons, capB, capC, and capA, in this order of arrangement. Minicell analysis revealed that these cistrons produce proteins of 44, 16, and 46 kilodaltons, respectively. The complete nucleotide sequence of 3,244 base pairs covering the whole cap region was determined and revealed the existence of the three open reading frames of capB (397 amino acid residues; molecular weight, 44,872), capC (149 amino acid residues; molecular weight, 16,522), and capA (411 amino acid residues; molecular weight, 46,420) arranged in the order predicted by complementation tests. These three cistrons were all transcribed in the same direction from promoters unique to each cistron. Judging from the predicted amino acid sequence of the three proteins and from their localization and their sensitivity to various physicochemical treatments, they appeared to be membrane-associated enzymes mediating the polymerization of D-glutamic acid via the membrane. Capsular peptides immunologically identical to that of B. anthracis were found in B. subtilis, B. megaterium, and B. licheniformis, but no sequence homologous to the cap region was found in any of these bacilli other than B. anthracis. Using strains of B. anthracis with or without insertional inactivation of the cap region, we found that the capsule of B. anthracis conferred strong resistance to phagocytosis upon the bacterial host.
机译:通过对各种体外构建的突变体进行遗传互补测试,这些突变体的帽区域具有突变(这对于封装在炭疽芽胞杆菌中至关重要),我们按此排列顺序确定了三个顺反子,capB,capC和capA。微型细胞分析表明,这些顺反子分别产生44、16和46千道尔顿的蛋白质。确定了覆盖整个帽区域的3,244个碱基对的完整核苷酸序列,并揭示了capB(397个氨基酸残基;分子量为44,872),capC(149个氨基酸残基;分子量为16,522)的三个开放阅读框的存在。 )和capA(411个氨基酸残基;分子量46,420)按互补性测试预测的顺序排列。这三个顺反子均从每个顺反子独特的启动子沿相同方向转录。从这三种蛋白的预测氨基酸序列及其定位和对各种理化处理的敏感性来看,它们似乎是与膜相关的酶,介导通过膜的D-谷氨酸聚合。在枯草芽孢杆菌,巨大芽孢杆菌和地衣芽孢杆菌中发现了与炭疽芽孢杆菌在免疫学上相同的荚膜肽,但是在除炭疽芽孢杆菌之外的任何这些细菌中均未发现与帽区域同源的序列。使用带有或不带帽盖插入失活的炭疽芽孢杆菌菌株,我们发现炭疽芽孢杆菌的胶囊赋予细菌宿主强烈的抗吞噬能力。

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