Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis
Cellular production of tryptophan is metabolically expensive and tightly regulated. The small Bacillus subtilis zinc binding Anti-TRAP protein (AT), which is the product of the yczA/rtpA gene, is upregulated in response to accumulating levels of uncharged tRNATrp through a T-box antitermination mech...
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Elsevier
2024-12-01
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| Series: | Journal of Structural Biology: X |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2590152424000084 |
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| author | Craig A. McElroy Elihu C. Ihms Deepak Kumar Yadav Melody L. Holmquist Vibhuti Wadhwa Vicki H. Wysocki Paul Gollnick Mark P. Foster |
| author_facet | Craig A. McElroy Elihu C. Ihms Deepak Kumar Yadav Melody L. Holmquist Vibhuti Wadhwa Vicki H. Wysocki Paul Gollnick Mark P. Foster |
| author_sort | Craig A. McElroy |
| collection | DOAJ |
| description | Cellular production of tryptophan is metabolically expensive and tightly regulated. The small Bacillus subtilis zinc binding Anti-TRAP protein (AT), which is the product of the yczA/rtpA gene, is upregulated in response to accumulating levels of uncharged tRNATrp through a T-box antitermination mechanism. AT binds to the undecameric axially symmetric ring-shaped protein TRAP (trp RNA Binding Attenuation Protein), thereby preventing it from binding to the trp leader RNA. This reverses the inhibitory effect of TRAP on transcription and translation of the trp operon. AT principally adopts two symmetric oligomeric states, a trimer (AT3) featuring three-fold axial symmetry or a dodecamer (AT12) comprising a tetrahedral assembly of trimers, whereas only the trimeric form binds and inhibits TRAP. We apply native mass spectrometry (nMS) and small-angle x-ray scattering (SAXS), together with analytical ultracentrifugation (AUC) to monitor the pH and concentration-dependent equilibrium between the trimeric and dodecameric structural forms of AT. In addition, we use solution nuclear magnetic resonance (NMR) spectroscopy to determine the solution structure of AT3, while heteronuclear 15N relaxation measurements on both oligomeric forms of AT provide insights into the dynamic properties of binding-active AT3 and binding-inactive AT12, with implications for TRAP binding and inhibition. |
| format | Article |
| id | doaj-art-97ca08ddeeaa4d86bfaef1e8ed2b514e |
| institution | DOAJ |
| issn | 2590-1524 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Structural Biology: X |
| spelling | doaj-art-97ca08ddeeaa4d86bfaef1e8ed2b514e2025-08-20T02:50:27ZengElsevierJournal of Structural Biology: X2590-15242024-12-011010010310.1016/j.yjsbx.2024.100103Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilisCraig A. McElroy0Elihu C. Ihms1Deepak Kumar Yadav2Melody L. Holmquist3Vibhuti Wadhwa4Vicki H. Wysocki5Paul Gollnick6Mark P. Foster7Ohio State Biochemistry Program, USA; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USADepartment of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; Biophysics Program, USADepartment of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USAOhio State Biochemistry Program, USA; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USADepartment of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USADepartment of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; National Resource for Native MS-Guided Structural Biology, USADepartment of Biological Sciences, State University of New York, Buffalo, NY 14260, USAOhio State Biochemistry Program, USA; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; Biophysics Program, USA; Corresponding author at: Department of Chemistry and Biochemistry, USA.Cellular production of tryptophan is metabolically expensive and tightly regulated. The small Bacillus subtilis zinc binding Anti-TRAP protein (AT), which is the product of the yczA/rtpA gene, is upregulated in response to accumulating levels of uncharged tRNATrp through a T-box antitermination mechanism. AT binds to the undecameric axially symmetric ring-shaped protein TRAP (trp RNA Binding Attenuation Protein), thereby preventing it from binding to the trp leader RNA. This reverses the inhibitory effect of TRAP on transcription and translation of the trp operon. AT principally adopts two symmetric oligomeric states, a trimer (AT3) featuring three-fold axial symmetry or a dodecamer (AT12) comprising a tetrahedral assembly of trimers, whereas only the trimeric form binds and inhibits TRAP. We apply native mass spectrometry (nMS) and small-angle x-ray scattering (SAXS), together with analytical ultracentrifugation (AUC) to monitor the pH and concentration-dependent equilibrium between the trimeric and dodecameric structural forms of AT. In addition, we use solution nuclear magnetic resonance (NMR) spectroscopy to determine the solution structure of AT3, while heteronuclear 15N relaxation measurements on both oligomeric forms of AT provide insights into the dynamic properties of binding-active AT3 and binding-inactive AT12, with implications for TRAP binding and inhibition.http://www.sciencedirect.com/science/article/pii/S2590152424000084Homo-oligomerizationNMR SpectroscopySAXSAUCNative MS |
| spellingShingle | Craig A. McElroy Elihu C. Ihms Deepak Kumar Yadav Melody L. Holmquist Vibhuti Wadhwa Vicki H. Wysocki Paul Gollnick Mark P. Foster Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis Journal of Structural Biology: X Homo-oligomerization NMR Spectroscopy SAXS AUC Native MS |
| title | Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis |
| title_full | Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis |
| title_fullStr | Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis |
| title_full_unstemmed | Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis |
| title_short | Solution structure, dynamics and tetrahedral assembly of Anti-TRAP, a homo-trimeric triskelion-shaped regulator of tryptophan biosynthesis in Bacillus subtilis |
| title_sort | solution structure dynamics and tetrahedral assembly of anti trap a homo trimeric triskelion shaped regulator of tryptophan biosynthesis in bacillus subtilis |
| topic | Homo-oligomerization NMR Spectroscopy SAXS AUC Native MS |
| url | http://www.sciencedirect.com/science/article/pii/S2590152424000084 |
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