EXPRESSION OF THE O9 POLYSACCHARIDE OF ESCHERICHIA-COLI - SEQUENCING OF THE ESCHERICHIA-COLI O9 RFB GENE-CLUSTER, CHARACTERIZATION OF MANNOSYL TRANSFERASES, AND EVIDENCE FOR AN ATP-BINDING CASSETTE TRANSPORT-SYSTEM
EXPRESSION OF THE O9 POLYSACCHARIDE OF ESCHERICHIA-COLI - SEQUENCING OF THE ESCHERICHIA-COLI O9 RFB GENE-CLUSTER, CHARACTERIZATION OF MANNOSYL TRANSFERASES, AND EVIDENCE FOR AN ATP-BINDING CASSETTE TRANSPORT-SYSTEM
复制标题
DOI：
10.1128/jb.177.8.2178-2187.1995
复制
发表时间：
1995-04-01
影响因子：
3.2
通讯作者：
JANN, K
中科院分区：
文献类型：
--
作者：
KIDO, N;TORGOV, VI;JANN, K
大肠杆菌O 9的rfb基因簇指导O 9特异性多糖的合成，所述多糖具有结构-->2-α-Man-（1-->2）-α-Man-（1-->2）-α-Man-（1-->3）-α-Man-（1-->3）-α-Man-（1-->。急诊coliO 9 rfb簇进行了序列测定，除先前描述的rfbK和rfbM外，还鉴定了6个基因。它们对应于编码261、431、708、815、381和274个氨基酸的多肽的六个开放阅读框（ORF）。它们的转录方向都与his操纵子相反。rfb与他的rfb之间没有发现基因。较高的G+C含量表明E. coliO 9 rfb的进化独立于其他大肠杆菌的rfb簇。大肠杆菌菌株和来自志贺氏菌和沙门氏菌属的菌株。缺失诱变，结合从突变体分离的膜中O 9甘露聚糖的体外合成分析，表明三个基因（称为mtfA，-B和-C，分别编码815，381和274个氨基酸的多肽）指导α-甘露糖基转移酶。MtfC（来自ORF 274），第一个甘露糖基转移酶，将甘露糖转移到内源性受体。它主要依赖于一个功能性rfe基因（该基因指导内源受体的合成）并启动多糖链的生长。MtfB（来自ORF 381）然后将两个甘露糖转移到先前甘露糖的3位，并且MtfA（来自ORF 815）将三个甘露糖转移到2位。进一步的链生长仅需要两种转移酶MtfA和MtfB。因此，所需的转移酶比重复单元中的糖的数量少。ORF 261和ORF 431蛋白的预测氨基酸序列的分析表明，它们作为ATP结合盒转运系统的组分起作用。一个可能的相关性的聚合机制和膜易位的产品的模式进行了讨论。
The rfb gene cluster of Escherichia coli O9 directs the synthesis of the O9-specific polysaccharide which has the structure -->2-alpha-Man-(1-->2)-alpha-Man-(1-->2)-alpha-Man-(1-->3)-alpha-Man-(1-->3)-alpha-Man-(1-->. The E. coli O9 rfb cluster has been sequenced, and six genes, in addition to the previously described rfbK and rfbM, were identified. They correspond to six open reading frames (ORFs) encoding polypeptides of 261, 431, 708, 815, 381, and 274 amino acids. They are all transcribed in the counter direction to those of the his operon. No gene was found between rfb and his. A higher G+C content indicated that E. coli O9 rfb evolved independently of the rfb clusters from other E. coli strains and from Shigella and Salmonella spp. Deletion mutagenesis, in combination with analysis of the in vitro synthesis of the O9 mannan in membranes isolated from the mutants, showed that three genes (termed mtfA, -B, and -C, encoding polypeptides of 815, 381, and 274 amino acids, respectively) directed alpha-mannosyl transferases. MtfC (from ORF274), the first mannosyl transferase, transfers a mannose to the endogenous acceptor. It critically depended on a functional rfe gene (which directs the synthesis of the endogenous acceptor) and initiates the growth of the polysaccharide chain. MtfB (from ORF381) then transfers two mannoses into the 3 position of the previous mannose, and MtfA (from ORF815) transfers three mannoses into the 2 position. Further chain growth needs only the two transferases MtfA and MtfB. Thus, there are fewer transferases needed than the number of sugars in the repeating unit. Analysis of the predicted amino acid sequence of the ORF261 and ORF431 proteins indicated that they function as components of an ATP binding cassette transport system. A possible correlation between the mechanism of polymerization and mode of membrane translocation of the products is discussed.