scholarly journals Glycosylation reduces the glycan-independent immunomodulatory effect of recombinant Orysata lectin in Drosophila S2 cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Pengyu Chen ◽  
Kristof De Schutter ◽  
Sonia Serna ◽  
Simin Chen ◽  
Qun Yang ◽  
...  
Keyword(s):  
Immune Responses ◽  
Protein Function ◽  
Expression System ◽  
Carbohydrate Binding ◽  
S2 Cells ◽  
Post Translational Modification ◽  
Recombinant Production ◽  
Mannose Binding ◽  
Binding Lectin ◽  
Selection Of

AbstractSeveral plant lectins, or carbohydrate-binding proteins, interact with glycan moieties on the surface of immune cells, thereby influencing the immune response of these cells. Orysata, a mannose-binding lectin from rice, has been reported to exert immunomodulatory activities on insect cells. While the natural lectin is non-glycosylated, recombinant Orysata produced in the yeast Pichia pastoris (YOry) is modified with a hyper-mannosylated N-glycan. Since it is unclear whether this glycosylation can affect the YOry activity, non-glycosylated rOrysata was produced in Escherichia coli (BOry). In a comparative analysis, both recombinant Orysata proteins were tested for their carbohydrate specificity on a glycan array, followed by the investigation of the carbohydrate-dependent agglutination of red blood cells (RBCs) and the carbohydrate-independent immune responses in Drosophila melanogaster S2 cells. Although YOry and BOry showed a similar carbohydrate-binding profiles, lower concentration of BOry were sufficient for the agglutination of RBCs and BOry induced stronger immune responses in S2 cells. The data are discussed in relation to different hypotheses explaining the weaker responses of glycosylated YOry. In conclusion, these observations contribute to the understanding how post-translational modification can affect protein function, and provide guidance in the selection of the proper expression system for the recombinant production of lectins.

scholarly journals Induction of Recombinant Lectin Expression by an Artificially Constructed Tandem Repeat Structure: A Case Study Using Bryopsis plumosa Mannose-Binding Lectin

Biomolecules ◽  
2018 ◽  
Vol 8 (4) ◽  
pp. 146 ◽  
Author(s):  
Hyun-Ju Hwang ◽  
Jin-Woo Han ◽  
Hancheol Jeon ◽  
Jong Han
Keyword(s):  
Tandem Repeat ◽  
Large Scale ◽  
Tandem Repeats ◽  
Expression System ◽  
Mannose Binding Lectin ◽  
Repeat Structure ◽  
Tandem Repeat Sequences ◽  
Mannose Binding ◽  
Bryopsis Plumosa ◽  
Binding Lectin

Lectin is an important protein in medical and pharmacological applications. Impurities in lectin derived from natural sources and the generation of inactive proteins by recombinant technology are major obstacles for the use of lectins. Expressing recombinant lectin with a tandem repeat structure can potentially overcome these problems, but few studies have systematically examined this possibility. This was investigated in the present study using three distinct forms of recombinant mannose-binding lectin from Bryopsis plumosa (BPL2)—i.e., the monomer (rD1BPL2), as well as the dimer (rD2BPL2), and tetramer (rD4BPL2) arranged as tandem repeats. The concentration of the inducer molecule isopropyl β-D-1-thiogalactopyranoside and the induction time had no effect on the efficiency of the expression of each construct. Of the tested constructs, only rD4BPL2 showed hemagglutination activity towards horse erythrocytes; the activity of towards the former was 64 times higher than that of native BPL2. Recombinant and native BPL2 showed differences in carbohydrate specificity; the activity of rD4BPL2 was inhibited by the glycoprotein fetuin, whereas that of native BPL2 was also inhibited by d-mannose. Our results indicate that expression as tandem repeat sequences can increase the efficiency of lectin production on a large scale using a bacterial expression system.

A novel mannose-binding lectin from Liparis nervosa with anti-fungal and anti-tumor activities

2020 ◽  
Vol 52 (10) ◽  
pp. 1081-1092
Author(s):  
Na Jiang ◽  
Yuqing Wang ◽  
Jing Zhou ◽  
Ruxiao Zheng ◽  
Xiao Yuan ◽  
...  
Keyword(s):  
Gel Filtration ◽  
Sclerotium Rolfsii ◽  
Exchange Chromatography ◽  
Mannose Binding Lectin ◽  
Carbohydrate Binding ◽  
Plant Lectins ◽  
Mannose Binding ◽  
Anti Fungal ◽  
Binding Lectin

Abstract Plant lectins are carbohydrate-binding proteins with nonimmune origin, which can reversibly bind with carbohydrates, agglutinate cells, and precipitate polysaccharides and glycoconjugates. Plant lectins have attracted much attention for their anti-virus, anti-proliferation, and pro-apoptosis properties. Thus the exploration of new lectins has received special attention. Here we purified a mannose-binding lectin from the rhizomes of Liparis nervosa by ion exchange chromatography on DEAE-Sepharose, affinity chromatography on Mannose-Sepharose 4B, and gel filtration chromatography on Sephacryl S-100. The purified L. nervosa lectin (LNL) was identified to be a monomeric protein with a molecular mass of 13 kDa. LNL exhibited hemagglutinating activity towards rabbit erythrocytes, and its activity could be strongly inhibited by D-mannose, N-acetyl glucosamine and thyroglobulin. In vitro experiments showed that LNL exhibited a comparable anti-fungal activity against Piricularia oryzae (Cavara), Bipolaris maydis, Fusarium graminearum, and Sclerotium rolfsii, and anti-proliferation activity against tumor cells by inducing apoptosis. The full-length cDNA sequence of LNL is 715 bp in length and contains a 525 bp open reading frame (ORF) encoding a 110-residue mature protein. It was predicted to have three mannose-binding conserved motifs ‘QXDXNXVXY’. The binding pattern of LNL was further revealed by homology modeling and molecular docking. We demonstrated that LNL is not only a potential therapeutic candidate against tumor but also a new anti-fungal agent.

Purification of Recombinant Mouse C-Reactive Protein from Pichia Pastoris GS115 by Nickel Chelating Sepharose Fast-Flow Affinity Chromatography and P-Aminophenyl Phosphoryl Choline Agarose Resin Affinity Chromatography in Tandem

2021 ◽  
Author(s):  
Bin Cheng ◽  
Di Wu ◽  
Ke Wu ◽  
Xiao-Ping Huang ◽  
Jian-Min Lv ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Affinity Chromatography ◽  
Protein Function ◽  
Recombinant Expression ◽  
Expression System ◽  
Post Translational Modification ◽  
C Reactive Protein ◽  
Reactive Protein ◽  
Phosphoryl Choline ◽  
Fast Flow

Abstract C-reactive protein (CRP) is a circulating marker of inflammation yet with ill-defined biological functions. This is partly due to the uncharacterized activities of endogenous CRP in mice, the major animal model used to define protein function. The hurdles for purification and characterization of mouse CRP are its low circulating levels and the lack of specific antibodies. To clear these hurdles, here we developed an efficient expression system by constructing recombinant Pichia pastoris cells for secretion of native conformation mouse CRP. The recombinant expression of mouse CRP in Escherichia coli failed to yield sufficient amount of native protein, reflecting the importance of post-translational modification of glycosylation in aiding proper folding. By contrast, sufficient amount of native mouse CRP was successfully purified from P. pastoris. Preliminary purification was performed by Nickel Chelating Sepharose Fast-Flow affinity chromatography with 6 × His tags attached to the protein. Subsequently, p-Aminophenyl Phosphoryl Choline Agarose resin affinity chromatography was used for tandem purification. The purified mouse CRP showed native pentamer and capabilities of PC binding. Moreover, the 6 × His tag provides a convenient tool for detecting the interactions of mouse CRP with ligands.

scholarly journals Relationship between Cognition, levels of PTX-3, MBL and their polymorphisms: A systematic review

2020 ◽  
Vol 9 (10) ◽  
pp. e8749109215
Author(s):  
Tatiana Lins Carvalho ◽  
Taciana Furtado de Mendonça Belmont ◽  
Andreia Soares da Silva ◽  
Kleyton Palmeira do Ó ◽  
Lucas de Lucena Simões e Silva ◽  
...  
Keyword(s):  
Systematic Review ◽  
Cognitive Processes ◽  
Genetic Influences ◽  
Pentraxin 3 ◽  
Eligibility Criteria ◽  
Mannose Binding ◽  
The Central Nervous System ◽  
Two Stages ◽  
Binding Lectin ◽  
Selection Of

Introduction: Studies have suggested that pentraxin 3 (PTX3) and Mannose Binding Lectin (MBL) may interfere with cognitive processes. Objective: Identify data reported in the literature involving cognition, PTX3 and MBL. Materials and Methods: The research was done in five databases and the selection of studies was performed in two stages. The first involved review of titles and abstracts. Then the full articles were read and those that did not meet the eligibility criteria were excluded. Results: A total of 3,097 titles and abstracts were selected, but 3,089 were excluded. Finally, 8 articles were included in the review. The articles pointed out that high levels of PTX-3 could be predictors of cognitive impairment while high levels of MBL could have a protective effect on cognition. Conclusion: The current studies are still contradictory and inconclusive, but they lead us to reflect on possible genetic influences of innate immunity in the Central Nervous System. Further research involving the effects of PTX-3 and MBL and its variants on cognition are necessary.

scholarly journals Immune responses upon experimental Erysipelothrix rhusiopathiae infection of naïve and vaccinated chickens

2020 ◽  
Vol 51 (1) ◽  
Author(s):  
Eva Wattrang ◽  
Helena Eriksson ◽  
Tomas Jinnerot ◽  
Maria Persson ◽  
Elisabeth Bagge ◽  
...  
Keyword(s):  
Immune Responses ◽  
Acute Phase ◽  
Serum Levels ◽  
Mannose Binding Lectin ◽  
Erysipelothrix Rhusiopathiae ◽  
Naive Group ◽  
Mannose Binding ◽  
Leukocyte Counts ◽  
Blood Leukocyte ◽  
Binding Lectin

Abstract Erysipelas, a disease caused by Erysipelothrix rhusiopathiae (ER), is an increasing problem in laying hens housed in cage-free systems. This study aimed to monitor immune responses during ER infection of naïve chickens and chickens vaccinated intra muscularly with a commercial inactivated ER vaccine. Chickens were infected intra muscularly with ER at 30 days of age and blood leukocyte counts, serum levels of mannose binding lectin (MBL) and ER-specific IgY were monitored until the experiment was terminated at day 15 after infection. ER was detected in blood from more chickens and at higher bacterial counts in the naïve group (day 1: 1 of 7 chickens; day 3: 6 of 6 chickens) than in the vaccinated group (day 1: 0 of 7 chickens; day 3: 1 of 6 chickens). During the acute phase of infection transient increases in circulating heterophil numbers and serum MBL levels were detected in all ER infected chickens but these responses were prolonged in chickens from the naïve group compared to vaccinated chickens. Before infection IgY titers to ER in vaccinated chickens did not differ significantly from those of naïve chickens but vaccinated chickens showed significantly increased IgY titers to ER earlier after infection compared to chickens in the naïve group. In conclusion, the ER infection elicited prompt acute innate responses in all chickens. Vaccinated chickens did not have high IgY titers to ER prior to infection but did however show lower levels of bacteraemia and their acute immune responses were of shorter duration.

scholarly journals Proteomic Approaches to Dissect Host SUMOylation during Innate Antiviral Immune Responses

Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 528
Author(s):  
Marie Lork ◽  
Gauthier Lieber ◽  
Benjamin G. Hale
Keyword(s):  
Immune Responses ◽  
Large Scale ◽  
Future Research ◽  
Virus Infections ◽  
Restriction Factors ◽  
Post Translational Modification ◽  
Innate Defense ◽  
Cellular Stresses ◽  
Interferon Responses ◽  
Selection Of

SUMOylation is a highly dynamic ubiquitin-like post-translational modification that is essential for cells to respond to and resolve various genotoxic and proteotoxic stresses. Virus infections also constitute a considerable stress scenario for cells, and recent research has started to uncover the diverse roles of SUMOylation in regulating virus replication, not least by impacting antiviral defenses. Here, we review some of the key findings of this virus-host interplay, and discuss the increasingly important contribution that large-scale, unbiased, proteomic methodologies are making to discoveries in this field. We highlight the latest proteomic technologies that have been specifically developed to understand SUMOylation dynamics in response to cellular stresses, and comment on how these techniques might be best applied to dissect the biology of SUMOylation during innate immunity. Furthermore, we showcase a selection of studies that have already used SUMO proteomics to reveal novel aspects of host innate defense against viruses, such as functional cross-talk between SUMO proteins and other ubiquitin-like modifiers, viral antagonism of SUMO-modified antiviral restriction factors, and an infection-triggered SUMO-switch that releases endogenous retroelement RNAs to stimulate antiviral interferon responses. Future research in this area has the potential to provide new and diverse mechanistic insights into host immune defenses.

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