Genus-specific protein binding to the large clusters of DNA repeats (short regularly spaced repeats) present in Sulfolobus genomes
Genus-specific protein binding to the large clusters of DNA repeats (short regularly spaced repeats) present in Sulfolobus genomes
复制标题
与硫球菌基因组中存在的大量 DNA 重复序列（间隔规则的短重复序列）结合的基因特异性蛋白质
DOI：
10.1128/jb.185.8.2410-2417.2003
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发表时间：
2003-04-01
影响因子：
3.2
通讯作者：
Garrett, RA
中科院分区：
文献类型：
--
作者：
Peng, X;Brügger, M;Garrett, RA
短规则间隔重复序列（SRSRs）在古细菌染色体中以多个大簇出现，在一些古细菌共轭质粒和细菌染色体中以较小的簇出现。序列、大小和重复序列的间隔在一个集群内通常是恒定的，但在集群之间是不同的。对于crenarchaeon Sulfolobus solfataricus P2，基因组中的重复序列主要分为两个密切相关的序列家族，它们排列在7个簇中，共包含441个重复序列，约占基因组的1%。Sulfolobus共轭质粒pNOB8包含一个由6个重复序列组成的小簇，这些重复序列与S. solfataricus染色体上的一个重复变异序列相同。扩增了pNOB8序列的重复序列，并测试了其与葡萄细胞提取物的蛋白结合。从细胞提取物中纯化出17.5 kda的srsr结合蛋白并进行测序。该蛋白被N端修饰，对应于SSO454，一个先前未分配功能的开放阅读框。它与携带双重复序列和单重复序列的DNA片段特异性结合，结合在重复结构的一侧，并在DNA结构的另一侧产生开口。它还能识别出葡萄球菌重复序列的两个主要家族。在大肠杆菌中表达的重组蛋白与天然的SRSR重复序列具有相同的结合特性。SSO454蛋白具有三方内部重复结构，与螺旋-螺旋-螺旋dna结合基序具有良好的序列匹配。尽管这一假定的基序与其他古细菌蛋白相同，但SSO454的同源物仅在Sulfolobus属和与之密切相关的Acidianus属的物种中被检测到。我们推断，该属特异性蛋白诱导每个DNA重复中心的结构打开，从而为另一种蛋白质（可能是更保守的蛋白质）产生结合位点，这一过程可能对SRSR簇的高阶结构至关重要。
Short regularly spaced repeats (SRSRs) occur in multiple large clusters in archaeal chromosomes and as smaller clusters in some archaeal conjugative plasmids and bacterial chromosomes. The sequence, size, and spacing of the repeats are generally constant within a cluster but vary between clusters. For the crenarchaeon Sulfolobus solfataricus P2, the repeats in the genome fall mainly into two closely related sequence families that are arranged in seven clusters containing a total of 441 repeats which constitute ca. 1% of the genome. The Sulfolobus conjugative plasmid pNOB8 contains a small cluster of six repeats that are identical in sequence to one of the repeat variants in the S. solfataricus chromosome. Repeats from the pNOB8 cluster were amplified and tested for protein binding with cell extracts from S. solfataricus. A 17.5-kDa SRSR-binding protein was purified from the cell extracts and sequenced. The protein is N terminally modified and corresponds to SSO454, an open reading frame of previously unassigned function. It binds specifically to DNA fragments carrying double and single repeat sequences, binding on one side of the repeat structure, and producing an opening of the opposite side of the DNA structure. It also recognizes both main families of repeat sequences in S. solfataricus. The recombinant protein, expressed in Escherichia coli, showed the same binding properties to the SRSR repeat as the native one. The SSO454 protein exhibits a tripartite internal repeat structure which yields a good sequence match with a helix-turn-helix DNA-binding motif. Although this putative motif is shared by other archaeal proteins, orthologs of SSO454 were only detected in species within the Sulfolobus genus and in the closely related Acidianus genus. We infer that the genus-specific protein induces an opening of the structure at the center of each DNA repeat and thereby produces a binding site for another protein, possibly a more conserved one, in a process that may be essential for higher-order stucturing of the SRSR clusters.