Relevant elements of a Maize γ-zein domain involved in protein body biogenesis
The N-terminal proline-rich domain of γ-zein (Zera) plays an important role in protein body (PB) formation not only in the original host (maize seeds) but in a broad spectrum of eukaryotic cells. However, the elements within the Zera sequence that are involved in the biogenesis of PBs have not been...
| Autores: | , , , , , |
|---|---|
| Tipo de recurso: | artículo |
| Fecha de publicación: | 2010 |
| País: | España |
| Institución: | Universitat Autònoma de Barcelona |
| Repositorio: | Dipòsit Digital de Documents de la UAB |
| Idioma: | inglés |
| OAI Identifier: | oai:ddd.uab.cat:217109 |
| Acceso en línea: | https://ddd.uab.cat/record/217109 https://dx.doi.org/urn:doi:10.1074/jbc.M110.116285 |
| Access Level: | acceso abierto |
| Palabra clave: | Amino acid sequence motifs Cysteine residues Fluorescent protein Immunoelectron microscopy Molecular dynamics simulations Nicotiana benthamiana Proline-rich domains Site directed mutagenesis |
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oai:ddd.uab.cat:217109 |
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ES |
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España |
| spelling |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis Llop-Tous, Immaculada Madurga, Sergio Giralt, Ernest|||0000-0001-8381-1797 Marzabal, Pablo Torrent, Margarita Ludevid, M. Dolors Amino acid sequence motifs Cysteine residues Fluorescent protein Immunoelectron microscopy Molecular dynamics simulations Nicotiana benthamiana Proline-rich domains Site directed mutagenesis The N-terminal proline-rich domain of γ-zein (Zera) plays an important role in protein body (PB) formation not only in the original host (maize seeds) but in a broad spectrum of eukaryotic cells. However, the elements within the Zera sequence that are involved in the biogenesis of PBs have not been clearly identified. Here, we focused on amino acid sequence motifs that could be involved in Zera oligomerization, leading to PB-like structures in Nicotiana benthamiana leaves. By using fusions of Zera with fluorescent proteins, we found that the lack of the repeat region (PPPVHL) of Zera resulted in the secretion of the fusion protein but that this repeat by itself did not form PBs. Although the repeat region containing eight units was the most efficient for Zera self-assembly, shorter repeats of 4-6 units still formed small multimers. Based on site-directed mutagenesis of Zera cysteine residues and analysis of multimer formation, we conclude that the two N-terminal Cys residues of Zera (Cys and Cys) are critical for oligomerization. Immunoelectron microscopy and confocal studies on PB development over time revealed that early, small, Zera-derived oligomers were sequestered in buds along the rough ER and that the mature size of the PBs could be attained by both cross-linking of preformed multimers and the incorporation of new chains of Zera fusions synthesized by active membrane-bound ribosomes. Based on these results and on the behavior of the Zera structure determined by molecular dynamics simulation studies, we propose a model of Zera-induced PB biogenesis. https://ddd.uab.cat/record/217109 https://dx.doi.org/urn:doi:10.1074/jbc.M110.116285 |
| title |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis |
| spellingShingle |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis Llop-Tous, Immaculada Amino acid sequence motifs Cysteine residues Fluorescent protein Immunoelectron microscopy Molecular dynamics simulations Nicotiana benthamiana Proline-rich domains Site directed mutagenesis |
| title_short |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis |
| title_full |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis |
| title_fullStr |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis |
| title_full_unstemmed |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis |
| title_sort |
Relevant elements of a Maize γ-zein domain involved in protein body biogenesis |
| author |
Llop-Tous, Immaculada |
| author_facet |
Llop-Tous, Immaculada Madurga, Sergio Giralt, Ernest|||0000-0001-8381-1797 Marzabal, Pablo Torrent, Margarita Ludevid, M. Dolors |
| author_role |
author |
| author2 |
Madurga, Sergio Giralt, Ernest|||0000-0001-8381-1797 Marzabal, Pablo Torrent, Margarita Ludevid, M. Dolors |
| author2_role |
author author author author author |
| topic |
Amino acid sequence motifs Cysteine residues Fluorescent protein Immunoelectron microscopy Molecular dynamics simulations Nicotiana benthamiana Proline-rich domains Site directed mutagenesis |
| topic_facet |
Amino acid sequence motifs Cysteine residues Fluorescent protein Immunoelectron microscopy Molecular dynamics simulations Nicotiana benthamiana Proline-rich domains Site directed mutagenesis |
| description |
The N-terminal proline-rich domain of γ-zein (Zera) plays an important role in protein body (PB) formation not only in the original host (maize seeds) but in a broad spectrum of eukaryotic cells. However, the elements within the Zera sequence that are involved in the biogenesis of PBs have not been clearly identified. Here, we focused on amino acid sequence motifs that could be involved in Zera oligomerization, leading to PB-like structures in Nicotiana benthamiana leaves. By using fusions of Zera with fluorescent proteins, we found that the lack of the repeat region (PPPVHL) of Zera resulted in the secretion of the fusion protein but that this repeat by itself did not form PBs. Although the repeat region containing eight units was the most efficient for Zera self-assembly, shorter repeats of 4-6 units still formed small multimers. Based on site-directed mutagenesis of Zera cysteine residues and analysis of multimer formation, we conclude that the two N-terminal Cys residues of Zera (Cys and Cys) are critical for oligomerization. Immunoelectron microscopy and confocal studies on PB development over time revealed that early, small, Zera-derived oligomers were sequestered in buds along the rough ER and that the mature size of the PBs could be attained by both cross-linking of preformed multimers and the incorporation of new chains of Zera fusions synthesized by active membrane-bound ribosomes. Based on these results and on the behavior of the Zera structure determined by molecular dynamics simulation studies, we propose a model of Zera-induced PB biogenesis. |
| publishDate |
2010 |
| format |
article |
| url |
https://ddd.uab.cat/record/217109 https://dx.doi.org/urn:doi:10.1074/jbc.M110.116285 |
| language |
eng |
| eu_rights_str_mv |
openAccess |
| institution |
Universitat Autònoma de Barcelona |
| collection |
Dipòsit Digital de Documents de la UAB |
| reponame_str |
Dipòsit Digital de Documents de la UAB |
| instname_str |
Universitat Autònoma de Barcelona |
| _version_ |
1878437861802377216 |
| publishDateSort |
2010 |
| author_browse |
Giralt, Ernest|||0000-0001-8381-1797 Llop-Tous, Immaculada Ludevid, M. Dolors Madurga, Sergio Marzabal, Pablo Torrent, Margarita |
| score |
6,9303427 |