A mycobacteriophage genomics approach to identify novel mycobacteriophage proteins with mycobactericidal properties
A mycobacteriophage genomics approach to identify novel mycobacteriophage proteins with mycobactericidal properties
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DOI：
10.1099/mic.0.000810
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发表时间：
2019-07-01
期刊：
影响因子：
2.8
通讯作者：
Sen, Ranjan
中科院分区：
文献类型：
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作者：
Singh, Shweta;Godavarthi, Sapna;Sen, Ranjan
对分枝杆菌特异的分枝杆菌噬菌体是能够引发杀菌反应的各种效应蛋白的来源。我们描述了一种基因组学方法结合生物信息学来鉴定表达后对分枝杆菌有毒的分枝杆菌噬菌体蛋白。对包含噬菌体基因组集合的基因组文库筛选能够杀死耻垢分枝杆菌菌株mc 2155的克隆。我们鉴定了四个独特的克隆：克隆45和12 N（来自分枝杆菌噬菌体D29）和克隆66和85（来自分枝杆菌噬菌体Che 12）。来自克隆66和45的基因产物分别被鉴定为Che 12噬菌体的Gp 49和D29噬菌体的Gp 34。另外两个克隆85和12 N的基因产物利用编码合成蛋白的新开放阅读框（ORF）。这四个克隆（克隆45、66、85和12 N）引起M.耻垢病和牛分枝杆菌表达。Gp 49和Gp 34的克隆也诱导大肠杆菌的生长缺陷，表明它们靶向保守的宿主机制。它们的表达诱导各种形态学变化，表明它们影响DNA复制和细胞分裂步骤。我们预测，Gp 34是一个Xis蛋白，需要在噬菌体DNA切除从细菌染色体。Gp 49被预测为具有DNA弯曲/扭曲性质的HTH基序。我们建议，这种方法是有用的，以确定新的噬菌体蛋白质所需的性能，而无需费力地表征个别的噬菌体。它具有通用性，可应用于其他细菌-噬菌体系统。我们推测，具有独特基因产物的几乎无限数量的细菌的存在可以为探索新的抗菌分子提供更便宜，危害更小的替代方案。
Mycobacteriophages that are specific to mycobacteria are sources of various effector proteins that are capable of eliciting bactericidal responses. We describe a genomics approach in combination with bioinformatics to identify mycobacteriophage proteins that are toxic to mycobacteria upon expression. A genomic library comprising phage genome collections was screened for clones capable of killing Mycobacterium smegmatis strain mc2155. We identified four unique clones: clones 45 and 12N (from the mycobacteriophage D29) and clones 66 and 85 (from the mycobacteriophage Che12). The gene products from clones 66 and 45 were identified as Gp49 of the Che12 phage and Gp34 of the D29 phage, respectively. The gene products of the other two clones, 85 and 12N, utilized novel open reading frames (ORFs) coding for synthetic proteins. These four clones (clones 45, 66, 85 and 12N) caused growth defects in M. smegmatis and Mycobacterium bovis upon expression. Clones with Gp49 and Gp34 also induced growth defects in Escherichia coli, indicating that they target conserved host machineries. Their expression induced various morphological changes, indicating that they affected DNA replication and cell division steps. We predicted that Gp34 is a Xis protein that is required in phage DNA excision from the bacterial chromosome. Gp49 is predicted to have an HTH motif with DNA-bending/twisting properties. We suggest that this methodology is useful to identify new phage proteins with the desired properties without laboriously characterizing the individual phages. It is universal and could be applied to other bacteria-phage systems. We speculate that the existence of a virtually unlimited number of phages with unique gene products could offer a cheaper and less hazardous alternative to explore new antimicrobial molecules.