Localization of Native Mms13 to the Magnetosome Chain of Magnetospirillum magneticum AMB-1 Using Immunogold Electron Microscopy, Immunofluorescence Microscopy and Biochemical Analysis
| العنوان: | Localization of Native Mms13 to the Magnetosome Chain of Magnetospirillum magneticum AMB-1 Using Immunogold Electron Microscopy, Immunofluorescence Microscopy and Biochemical Analysis |
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| المؤلفون: | Steven K. Lower, Concepcion Jimenez-Lopez, Brian H. Lower, Carmen Valverde-Tercedor, Eric Mumper, Zachery Oestreicher, Lumarie Pérez-Guzmán, Dennis A. Bazylinski, Nadia N. Casillas-Ituarte |
| المصدر: | Digibug. Repositorio Institucional de la Universidad de Granada instname Crystals; Volume 11; Issue 8; Pages: 874 Crystals, Vol 11, Iss 874, p 874 (2021) |
| بيانات النشر: | MDPI AG, 2021. |
| سنة النشر: | 2021 |
| مصطلحات موضوعية: | Biomineralization, magnetite, Magnetotactic bacteria, General Chemical Engineering, Magnetosome, Nanocrystal, 02 engineering and technology, Tandem mass spectrometry, Immunofluorescence, nanocrystal, Magnetite, Inorganic Chemistry, 03 medical and health sciences, Protein sequencing, Western blot, medicine, General Materials Science, bacteria, magnetosome, 030304 developmental biology, 0303 health sciences, Crystallography, Bacteria, medicine.diagnostic_test, Molecular mass, Chemistry, Proteins, Magnetotactic, magnetotactic, Immunogold labelling, biomineralization, 021001 nanoscience & nanotechnology, Condensed Matter Physics, Molecular biology, QD901-999, protein, TEM, 0210 nano-technology |
| الوصف: | Magnetotactic bacteria (MTB) biomineralize intracellular magnetite (Fe3O4 ) crystals surrounded by a magnetosome membrane (MM). The MM contains membrane-specific proteins that control Fe3O4 mineralization in MTB. Previous studies have demonstrated that Mms13 is a critical protein within the MM. Mms13 can be isolated from the MM fraction of Magnetospirillum magneticum AMB-1 and a Mms13 homolog, MamC, has been shown to control the size and shape of magnetite nanocrystals synthesized in-vitro. The objective of this study was to use several independent methods to definitively determine the localization of native Mms13 in M. magneticum AMB-1. Using Mms13-immunogold labeling and transmission electron microscopy (TEM), we found that Mms13 is localized to the magnetosome chain of M. magneticum AMB-1 cells. Mms13 was detected in direct contact with magnetite crystals or within the MM. Immunofluorescence detection of Mms13 in M. magneticum AMB-1 cells by confocal laser scanning microscopy (CLSM) showed Mms13 localization along the length of the magnetosome chain. Proteins contained within the MM were resolved by SDS-PAGE for Western blot analysis and LC-MS/MS (liquid chromatography with tandem mass spectrometry) protein sequencing. Using Anti-Mms13 antibody, a protein band with a molecular mass of ~14 kDa was detected in the MM fraction only. This polypeptide was digested with trypsin, sequenced by LC-MS/MS and identified as magnetosome protein Mms13. Peptides corresponding to the protein’s putative MM domain and catalytic domain were both identified by LC-MS/MS. Our results (Immunogold TEM, Immunofluorescence CLSM, Western blot, LC-MS/MS), combined with results from previous studies, demonstrate that Mms13 and homolog proteins MamC and Mam12, are localized to the magnetosome chain in MTB belonging to the class Alphaproteobacteria. Because of their shared localization in the MM and highly conserved amino acid sequences, it is likely that MamC, Mam12, and Mms13 share similar roles in the biomineralization of Fe3O4 nanocrystals. National Science Foundation, grant number EAR-2038207 EAR-1423939 Ministerio de Economía y Competitividad, SPAIN and Fondo Europeo de Desarrollo Regional, FEDER grant numbers CGL2010-18274 and CGL2013-46612 |
| وصف الملف: | application/pdf |
| تدمد: | 2073-4352 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::c6b250f34b956eabf23e0b696b8455b3 https://doi.org/10.3390/cryst11080874 |
| حقوق: | OPEN |
| رقم الأكسشن: | edsair.doi.dedup.....c6b250f34b956eabf23e0b696b8455b3 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 20734352 |
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