Assembly in vitro of Rhodococcus jostii RHA1 encapsulin and peroxidase DypB to form a nanocompartment
Assembly in vitro of Rhodococcus jostii RHA1 encapsulin and peroxidase DypB to form a nanocompartment
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DOI：
10.1111/febs.12234
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发表时间：
2013-05-01
期刊：
影响因子：
5.4
通讯作者：
Bugg, Timothy D. H.
中科院分区：
文献类型：
--
作者：
Rahmanpour, Rahman;Bugg, Timothy D. H.
红球菌 JostiiRHA1 过氧化物酶 DypB 最近被鉴定为细菌木质素过氧化物酶。 dypB 基因与编码封装蛋白的基因共转录，该基因已在 Thermotogamaritima 中显示组装形成 60 个亚基的纳米区室，并且 DypB 包含一个 C 端序列基序，被认为将蛋白质靶向封装蛋白纳米区室。 R.jostiiRHA1 封装蛋白在 R.jostii RHA1 中过表达，并纯化为高 Mr 组装体 (Mr>106)。通过在 pH3.0 下处理，纯化的纳米室可以被分解以形成低 Mr 物质，并重新组装以形成类似尺寸和形状的组件，如动态光散射所评估的。重组 DypB 可以在体外与单体封装蛋白组装，形成与仅封装蛋白纳米室相似尺寸的组件，如通过凝胶过滤评估的。通过硝化木质素紫外可见测定方法测定，与天然 DypB 相比，组装的复合物显示出每毫克 DypB 的木质素降解活性增强。复合物中测得的 8.6mol encapsulin/mol DypB 的化学计量与根据晶体结构预测的值 10 相似。结构化数字摘要encapsulin 和 encapsulin 通过蓝色本机页面结合（查看交互）encapsulin 和 encapsulin 通过动态光散射结合（查看交互）
RhodococcusjostiiRHA1 peroxidase DypB has been recently identified as a bacterial lignin peroxidase. The dypB gene is cotranscribed with a gene encoding an encapsulin protein, which has been shown in Thermotogamaritima to assemble to form a 60-subunit nanocompartment, and DypB contains a C-terminal sequence motif that is thought to target the protein to the encapsulin nanocompartment. R.jostiiRHA1 encapsulin protein was overexpressed in R.jostii RHA1, and purified as a high-Mr assembly (Mr>106). The purified nanocompartment could be disassembled to form a low-Mr species by treatment at pH3.0, and reassembled to form an assembly of similar size and shape, as assessed by dynamic light scattering. Recombinant DypB could be assembled invitro with monomeric encapsulin to form an assembly of similar size to the encapsulin-only nanocompartment, as assessed by gel filtration. The assembled complex showed enhanced lignin degradation activity per milligram of DypB present as compared with native DypB, as determined with a nitrated lignin UVvisible assay method. The measured stoichiometry of 8.6mol encapsulin/mol DypB in the complex was similar to the value of 10 predicted from the crystal structure.Structured digital abstractencapsulin and encapsulin bind by blue native page (View interaction)encapsulin and encapsulin bind by dynamic light scattering (View interaction)