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

Sousa, FD; Da Silva, BB; Furtado, GP; Carneiro, IDS; Lobo, MDP; Guan, Y; Guo, J; ... Moreira, RDA; + view all (2017) Frutapin, a lectin from Artocarpus incisa (breadfruit): cloning, expression and molecular insights. Bioscience Reports , 37 (4) , Article BSR20170969. 10.1042/BSR20170969. Green open access

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Abstract

Artocarpus incisa (breadfruit) seeds contain three different lectins (Frutalin, Frutapin 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. Frutapin (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 E. 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 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 simulation suggested FTP displays higher affinity for mannose than glucose. Cell studies revealed FTP was non-cytotoxic to cultured mouse fibroblast 3T3 cells below 0.5 mg/mL and 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.

Type: Article
Title: Frutapin, a lectin from Artocarpus incisa (breadfruit): cloning, expression and molecular insights
Location: England
Open access status: An open access version is available from UCL Discovery
DOI: 10.1042/BSR20170969
Publisher version: http://dx.doi.org/10.1042/BSR20170969
Language: English
Additional information: Copyright © 2017 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY) (http://creativecommons.org/licenses/by/4.0/).
Keywords: Biotechnology, computational biochemistry, protein-carbohydrate interactions
UCL classification: UCL
UCL > Provost and Vice Provost Offices
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Medicine
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Medicine > Inflammation
URI: https://discovery.ucl.ac.uk/id/eprint/1563654
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