权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Gas vesicle formation of haloarchaea: functions of the accessory Gvp proteins.

haloarchaea 的气体囊泡形成：辅助 Gvp 蛋白的功能。

基本信息

批准号：
406273130
负责人：
Professorin Dr. Felicitas Pfeifer
金额：
--
依托单位：
Arbeitsgruppe Microbiology and Archaea
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2018
资助国家：
德国
起止时间：
2017-12-31 至 2021-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/406273130?language=en
关键词：
Gas vesicle formation haloarchaea functions

项目摘要

Gas vesicles are intracellular nanostructures consisting of a proteinaceous wall surrounding a gas-filled space. GvpA and GvpC are the major constituents of this wall. GvpA forms antiparallel dimers that aggregate into ribs running as low-pitch helix perpendicular to the long axis. GvpC stabilizes the gas vesicle structure by attaching to the exterior surface. Ten additional Gvp proteins encoded by the early transcription unit gvpFGHIJKLM and the gvpACNO gene cluster are produced in minor amounts, and most of them are essential for gas-vesicle assembly. These accessory Gvp proteins might form protein-protein complexes required at the onset or during gas-vesicle formation. To investigate protein-protein interactions in haloarchaea we successfully adapted the split-GFP method. In addition, protein complexes were selected from haloarchaea using the cellulose-binding domain, CBD. The GvpM-GvpL interaction was confined to the N-terminal 25-aa of GvpM by split-GFP. Also, GvpH and GvpJ bind GvpM, and we propose to determine the binding sites of both in GvpM. If the binding sites of the three accessory Gvp overlap in GvpM, it is likely that they bind consecutively. A GvpM-CBD fusion selected GvpF, GvpH, GvpJ, GvpK, and GvpL from Hfx. volcanii transformants. These Gvp proteins appear to form a complex that might serve as nucleation site for the directed aggregation of GvpA at the onset of gas-vesicle assembly. We propose to study the FHJKLM-complex(es) to determine their size(s) and constituents. GvpF and GvpK are both constituents of this complex, and we propose to investigate whether they bind GvpM directly or whether other Gvp proteins mediate their interaction. Also, we will test the formation (and enlargement) of this complex in the presence of minor amounts of GvpA. GvpG and GvpI were not found so far. CBD-tagged GvpG and GvpI will be used to select putative binding partners. In addition, the interaction network of GvpA will be studied using fragments of this protein. Helix 1 will be tested as putative binding site between two ribs, and helix 2 might serves as regular anchor site of GvpC at the exterior surface of the gas vesicle wall. Also, the putative function of GvpN as AAA-ATPase will be investigated by substituting amino acids of the p-loop motif found in this protein.

气体囊泡是细胞内的纳米结构，其由围绕气体填充空间的蛋白质壁组成。GvpA和GvpC是该壁的主要成分。GvpA形成反平行二聚体，其聚集成作为垂直于长轴的低螺距螺旋运行的肋。GvpC通过附着到外表面来稳定气泡结构。由早期转录单位gvpFGHIJKLM和gvpACNO基因簇编码的另外10种Gvp蛋白质少量产生，并且它们中的大多数对于气体囊泡组装是必需的。这些辅助Gvp蛋白可能形成在发病时或在气体囊泡形成过程中所需的蛋白质-蛋白质复合物。为了研究盐古菌中的蛋白质-蛋白质相互作用，我们成功地采用了split-GFP方法。此外，使用纤维素结合结构域CBD从盐古菌中选择蛋白质复合物。通过split-GFP将GvpM-GvpL相互作用限制在GvpM的N-末端25-aa。此外，GvpH和GvpJ结合GvpM，我们建议确定两者在GvpM的结合位点。如果三个辅助Gvp的结合位点在GvpM中重叠，则它们可能连续结合。GvpM-CBD融合物从Hfx中选择GvpF、GvpH、GvpJ、GvpK和GvpL。volcanii转化体。这些Gvp蛋白似乎形成一个复杂的，可能作为成核位点的GvpA的直接聚集在开始的气体囊泡组装。我们建议研究FHJKLM-配合物，以确定它们的大小和组成。GvpF和GvpK都是这种复合物的组成部分，我们建议调查它们是否直接结合GvpM或其他Gvp蛋白介导它们的相互作用。此外，我们将测试在少量GvpA存在下该复合物的形成（和扩大）。GvpG和GvpI尚未发现。CBD标记的GvpG和GvpI将用于选择推定的结合配偶体。此外，GvpA的相互作用网络将使用该蛋白的片段进行研究。阻碍1将作为两个肋之间的假定结合位点进行测试，螺旋2可能作为GvpC在气囊壁外表面的规则锚位点。此外，GvpN作为AAA-ATP酶的推定功能将通过取代该蛋白质中发现的p-环基序的氨基酸来研究。