SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains.
The Nogo-B receptor (NgBR) is involved in oncogenic Ras signaling through directly binding to farnesylated Ras. It recruits farnesylated Ras to the non-lipid-raft membrane for interaction with downstream effectors. However, the cytosolic domain of NgBR itself is only partially folded. The lack of se...
| Main Authors: | , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2018-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC5773207?pdf=render |
| _version_ | 1818028917985902592 |
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| author | Joshua Holcomb Maysaa Doughan Nicholas Spellmon Brianne Lewis Emerson Perry Yingxue Zhang Lindsey Nico Junmei Wan Srinivas Chakravarthy Weifeng Shang Qing Miao Timothy Stemmler Zhe Yang |
| author_facet | Joshua Holcomb Maysaa Doughan Nicholas Spellmon Brianne Lewis Emerson Perry Yingxue Zhang Lindsey Nico Junmei Wan Srinivas Chakravarthy Weifeng Shang Qing Miao Timothy Stemmler Zhe Yang |
| author_sort | Joshua Holcomb |
| collection | DOAJ |
| description | The Nogo-B receptor (NgBR) is involved in oncogenic Ras signaling through directly binding to farnesylated Ras. It recruits farnesylated Ras to the non-lipid-raft membrane for interaction with downstream effectors. However, the cytosolic domain of NgBR itself is only partially folded. The lack of several conserved secondary structural elements makes this domain unlikely to form a complete farnesyl binding pocket. We find that inclusion of the extracellular and transmembrane domains that contain additional conserved residues to the cytosolic region results in a well folded protein with a similar size and shape to the E.coli cis-isoprenyl transferase (UPPs). Small Angle X-ray Scattering (SAXS) analysis reveals the radius of gyration (Rg) of our NgBR construct to be 18.2 Å with a maximum particle dimension (Dmax) of 61.0 Å. Ab initio shape modeling returns a globular molecular envelope with an estimated molecular weight of 23.0 kD closely correlated with the calculated molecular weight. Both Kratky plot and pair distribution function of NgBR scattering reveal a bell shaped peak which is characteristic of a single globularly folded protein. In addition, circular dichroism (CD) analysis reveals that our construct has the secondary structure contents similar to the UPPs. However, this result does not agree with the currently accepted topological orientation of NgBR which might partition this construct into three separate domains. This discrepancy suggests another possible NgBR topology and lends insight into a potential molecular basis of how NgBR facilitates farnesylated Ras recruitment. |
| first_indexed | 2024-12-10T05:11:25Z |
| format | Article |
| id | doaj.art-0d670d4ba5bf484b825e3347043161db |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-10T05:11:25Z |
| publishDate | 2018-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-0d670d4ba5bf484b825e3347043161db2022-12-22T02:01:05ZengPublic Library of Science (PLoS)PLoS ONE1932-62032018-01-01131e019137110.1371/journal.pone.0191371SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains.Joshua HolcombMaysaa DoughanNicholas SpellmonBrianne LewisEmerson PerryYingxue ZhangLindsey NicoJunmei WanSrinivas ChakravarthyWeifeng ShangQing MiaoTimothy StemmlerZhe YangThe Nogo-B receptor (NgBR) is involved in oncogenic Ras signaling through directly binding to farnesylated Ras. It recruits farnesylated Ras to the non-lipid-raft membrane for interaction with downstream effectors. However, the cytosolic domain of NgBR itself is only partially folded. The lack of several conserved secondary structural elements makes this domain unlikely to form a complete farnesyl binding pocket. We find that inclusion of the extracellular and transmembrane domains that contain additional conserved residues to the cytosolic region results in a well folded protein with a similar size and shape to the E.coli cis-isoprenyl transferase (UPPs). Small Angle X-ray Scattering (SAXS) analysis reveals the radius of gyration (Rg) of our NgBR construct to be 18.2 Å with a maximum particle dimension (Dmax) of 61.0 Å. Ab initio shape modeling returns a globular molecular envelope with an estimated molecular weight of 23.0 kD closely correlated with the calculated molecular weight. Both Kratky plot and pair distribution function of NgBR scattering reveal a bell shaped peak which is characteristic of a single globularly folded protein. In addition, circular dichroism (CD) analysis reveals that our construct has the secondary structure contents similar to the UPPs. However, this result does not agree with the currently accepted topological orientation of NgBR which might partition this construct into three separate domains. This discrepancy suggests another possible NgBR topology and lends insight into a potential molecular basis of how NgBR facilitates farnesylated Ras recruitment.http://europepmc.org/articles/PMC5773207?pdf=render |
| spellingShingle | Joshua Holcomb Maysaa Doughan Nicholas Spellmon Brianne Lewis Emerson Perry Yingxue Zhang Lindsey Nico Junmei Wan Srinivas Chakravarthy Weifeng Shang Qing Miao Timothy Stemmler Zhe Yang SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains. PLoS ONE |
| title | SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains. |
| title_full | SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains. |
| title_fullStr | SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains. |
| title_full_unstemmed | SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains. |
| title_short | SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains. |
| title_sort | saxs analysis of a soluble cytosolic ngbr construct including extracellular and transmembrane domains |
| url | http://europepmc.org/articles/PMC5773207?pdf=render |
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