Dataset Information

Frutapin, a lectin from Artocarpus incisa (breadfruit): cloning, expression and molecular insights.

ABSTRACT: Artocarpus incisa (breadfruit) seeds contain three different lectins (Frutalin, Frutapin (FTP) and Frutackin) with distinct carbohydrate specificities. The most abundant lectin is Frutalin, an ?-D-galactose-specific carbohydrate-binding glycoprotein with antitumour properties and potential for tumour biomarker discovery as already reported. FTP is the second most abundant, but proved difficult to purify with very low yields and contamination with Frutalin frustrating its characterization. Here, we report for the first time high-level production and isolation of biologically active recombinant FTP in Escherichia coli BL21, optimizing conditions with the best set yielding >40 mg/l culture of soluble active FTP. The minimal concentration for agglutination of red blood cells was 62.5 µg/ml of FTP, a process effectively inhibited by mannose. Apo-FTP, FTP-mannose and FTP-glucose crystals were obtained, and they diffracted X-rays to a resolution of 1.58 (P212121), 1.70 (P3121) and 1.60 (P3121) Å respectively. The best solution showed four monomers per asymmetric unit. Molecular dynamics (MD) simulation suggested that FTP displays higher affinity for mannose than glucose. Cell studies revealed that FTP was non-cytotoxic to cultured mouse fibroblast 3T3 cells below 0.5 mg/ml and was also capable of stimulating cell migration at 50 µg/ml. In conclusion, our optimized expression system allowed high amounts of correctly folded soluble FTP to be isolated. This recombinant bioactive lectin will now be tested in future studies for therapeutic potential; for example in wound healing and tissue regeneration.

SUBMITTER: de Sousa FD

PROVIDER: S-EPMC5520216 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Frutapin, a lectin from <i>Artocarpus incisa</i> (breadfruit): cloning, expression and molecular insights.

de Sousa Felipe Domingos FD da Silva Bruno Bezerra BB Furtado Gilvan Pessoa GP Carneiro Igor de Sa IS Lobo Marina Duarte Pinto MDP Guan Yiwei Y Guo Jingxu J Coker Alun R AR Lourenzoni Marcos Roberto MR Guedes Maria Izabel Florindo MIF Owen James S JS Abraham David J DJ Monteiro-Moreira Ana Cristina de Oliveira ACO Moreira Renato de Azevedo RA

Bioscience reports 20170721 4

<i>Artocarpus incisa</i> (breadfruit) seeds contain three different lectins (Frutalin, Frutapin (FTP) and Frutackin) with distinct carbohydrate specificities. The most abundant lectin is Frutalin, an α-D-galactose-specific carbohydrate-binding glycoprotein with antitumour properties and potential for tumour biomarker discovery as already reported. FTP is the second most abundant, but proved difficult to purify with very low yields and contamination with Frutalin frustrating its characterization. ...[more]

PMID: 28684550

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Project description:BackgroundBreadfruit (Artocarpus altilis) is a traditional staple tree crop throughout the tropics. Through interspecific grafting, a dwarf phenotype with over 50% reduction in plant height was identified when marang (Artocarpus odoratissimus) rootstocks were used. However, the molecular mechanism underlying the rootstock-induced breadfruit dwarfing is poorly understood.ResultsAn RNA-sequencing study of breadfruit scions at 22 months after grafting identified 5409 differentially expressed genes (DEGs) of which 2069 were upregulated and 3339 were downregulated in scion stems on marang rootstocks compared to those on self-graft. The DEGs were predominantly enriched for biological processes involved in carbon metabolism, cell wall organization, plant hormone signal transduction and redox homeostasis. The down-regulation of genes encoding vacuolar acid invertases and alkaline/neutral invertases, was consistent with the decreased activity of both enzymes, accompanying with a higher sucrose but lower glucose and fructose levels in the tissues. Key genes of biosynthetic pathways for amino acids, lipids and cell wall were down regulated, reflecting reduction of sucrose utilisation for stem growth on dwarfing rootstocks. Genes encoding sugar transporters, amino acid transporters, choline transporters, along with large number of potassium channels and aquaporin family members were down-regulated in scion stems on marang rootstocks. Lower activity of plasma membrane H+-ATPase, together with the predominance of genes encoding expansins, wall-associated receptor kinases and key enzymes for biosynthesis and re-modelling of cellulose, xyloglucans and pectins in down-regulated DGEs suggested impairment of cell expansion. Signalling pathways of auxin and gibberellin, along with strigolacton and brassinosteroid biosynthetic genes dominated the down-regulated DEGs. Phenylpropanoid pathway was enriched, with key lignin biosynthetic genes down-regulated, and flavonoid biosynthetic genes upregulated in scions on marang rootstocks. Signalling pathways of salicylic acid, jasmonic acid, ethylene and MAPK cascade were significantly enriched in the upregulated DEGs.ConclusionsRootstock-induced disruption in pathways regulating nutrient transport, sucrose utilisation, cell wall biosynthesis and networks of hormone transduction are proposed to impair cell expansion and stem elongation, leading to dwarf phenotype in breadfruit scions. The information provides opportunity to develop screening strategy for rootstock breeding and selection for breadfruit dwarfing.

Dataset Information

Frutapin, a lectin from Artocarpus incisa (breadfruit): cloning, expression and molecular insights.

Publications

Frutapin, a lectin from <i>Artocarpus incisa</i> (breadfruit): cloning, expression and molecular insights.

Similar Datasets

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