دورية أكاديمية
Recombinant N-glycosylation isoforms of Legume lectins: Production and purification from Nicotiana benthamiana leaves following RuBisCO depletion.
| العنوان: | Recombinant N-glycosylation isoforms of Legume lectins: Production and purification from Nicotiana benthamiana leaves following RuBisCO depletion. |
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| المؤلفون: | Bellande K; Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, Auzeville-Tolosane, France., Lalo A; Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, Auzeville-Tolosane, France., Ligat L; Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, Auzeville-Tolosane, France., Roujol D; Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, Auzeville-Tolosane, France., Jamet E; Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, Auzeville-Tolosane, France., Canut H; Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, Auzeville-Tolosane, France. Electronic address: canut@lrsv.ups-tlse.fr. |
| المصدر: | Plant physiology and biochemistry : PPB [Plant Physiol Biochem] 2020 Dec; Vol. 157, pp. 441-452. Date of Electronic Publication: 2020 Nov 05. |
| نوع المنشور: | Journal Article |
| اللغة: | English |
| بيانات الدورية: | Publisher: Elsevier Science Country of Publication: France NLM ID: 9882449 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1873-2690 (Electronic) Linking ISSN: 09819428 NLM ISO Abbreviation: Plant Physiol Biochem Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: Amsterdam : Elsevier Science Original Publication: Paris : Gauthier-Villars, c1987- |
| مواضيع طبية MeSH: | Ribulose-Bisphosphate Carboxylase*/metabolism, Fabaceae/*chemistry , Lectins/*biosynthesis , Nicotiana/*metabolism, Glycosylation ; Plant Leaves/metabolism ; Plants, Genetically Modified/metabolism ; Protein Isoforms/biosynthesis ; Recombinant Proteins/biosynthesis ; Nicotiana/genetics |
| مستخلص: | An efficient purification of recombinant proteins often requires a high ratio of recombinant to host proteins. In plants, Ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is the most abundant leaf protein, thus strongly impacting purification yield. Here, we describe a simple and robust purification procedure for recombinant proteins based on a differential precipitation of RuBisCO. In this context, four Legume lectin domains of Arabidopsis thaliana which belong to receptor-like kinases and cell wall proteins were produced from Nicotiana benthamiana leaves. The recombinant proteins exhibit a unique lectin domain consisting of around 250 amino acid residues with several predicted N-glycosylation sites and a six His-tag at the N-terminus. After ammonium sulphate precipitation of total soluble proteins, depletion of RuBisCO was obtained using citrate and succinate buffers during the salting-in step: this depletion was pH-dependent and the presence of di- or tri-carboxylic acids was required. The depleted protein extracts were then subjected to two chromatographic steps which were used in the negative mode to submit a protein fraction enriched as much as possible in recombinant lectin domains to a third chromatographic step (immobilized metal-ion chromatography). Three of the Legume lectin domains were purified near to homogeneity and revealed multiple N-glycosylation isoforms, particularly those from receptor-like kinases, which were characterised using specific lectins and deglycosylation enzymes. The production and purification of recombinant lectin domains will facilitate their biochemical characterisation in the context of cell-to-cell signalling and cell wall organisation. (Copyright © 2020 Elsevier Masson SAS. All rights reserved.) |
| فهرسة مساهمة: | Keywords: Arabidopsis thaliana; N-Glycosylation; Nicotiana benthamiana; Recombinant lectin; RuBisCO depletion |
| المشرفين على المادة: | 0 (Lectins) 0 (Protein Isoforms) 0 (Recombinant Proteins) EC 4.1.1.39 (Ribulose-Bisphosphate Carboxylase) |
| تواريخ الأحداث: | Date Created: 20201119 Date Completed: 20210127 Latest Revision: 20231213 |
| رمز التحديث: | 20240829 |
| DOI: | 10.1016/j.plaphy.2020.10.038 |
| PMID: | 33212361 |
| قاعدة البيانات: | MEDLINE |
Full Text Finder | تدمد: | 1873-2690 |
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| DOI: | 10.1016/j.plaphy.2020.10.038 |