Abstract
This study describes the molecular identification, biochemical characterization, and stabilization of three recombinant AlfA, AlfB, and AlfC fucosidases from Lacticaseibacillus rhamnosus INIA P603. Even though previous studies revealed the presence of fucosidase activity in L. rhamnosus extracts, the identification of the fucosidases, their physicochemical properties, and the substrate spectrum remained unknown. Although the presence of alfB is not common in strains of L. rhamnosus, fucosidases from L. rhamnosus INIA P603 were selected because this strain exhibited higher fucosidase activity in culture and the complete set of fucosidases. A high yield of purified recombinant AlfA, AlfB, and AlfC fucosidases was obtained (8, 12, and 18 mg, respectively). AlfA, AlfB, and AlfC showed their optimal activities at pH 5.0 and 4.0 at 60 °C, 40 °C, and 50 °C, respectively. Unlike 3-fucosyllactose, all three recombinant fucosidases were able to hydrolyze 2′-fucosyllactose (2′-FL), and their activities were improved through their immobilization on agarose supports. Nevertheless, immobilized AlfB exhibited the highest hydrolysis, releasing 39.6 µmol of fucose mg enzyme−1 min−1. Only the immobilized AlfB was able to synthetize 2′-FL. In conclusion, the enzymatic properties elucidated in this study support the potential ability of fucosidases from L. rhamnosus INIA P603 to hydrolyze fucosylated substrates as well as justifying interest for further research into AlfB for its application to catalyze the synthesis of fucosylated prebiotics.
Key points
• Few strains of L. rhamnosus exhibited alfB on their chromosomes.
• Fucosidases from L. rhamnosus INIA P603 were characterized and stabilized.
• Although all the fucosidases hydrolyzed 2′-FL, only AlfB transfucosylated lactose.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
Sequences of alfA, alfB, and alfC genes from Lacticaseibacillus rhamnosus INIA P603 were deposited in the GenBank under the following accession numbers as ON886905, ON886906, and ON886907, respectively. The authors declare that the data supporting the findings of this study are available within the article and its supplementary information files.
References
Becker DJ, Lowe JB (2003) Fucose: biosynthesis and biological function in mammals. Glycobiology 13:41R-53R. https://doi.org/10.1093/glycob/cwg054
Curiel JA, Rodríguez H, Acebrón I, Mancheño JM, De Las RB, Muñoz R (2009) Production and physicochemical properties of recombinant Lactobacillus plantarum tannase. J Agric Food Chem 57(14):6224–6230. https://doi.org/10.1021/jf901045s
Curiel JA, Peirotén Á, Landete JM, Ruiz de la Bastida A, Langa S, Arqués JL (2021) Architecture insight of bifidobacterial α-L-fucosidases. Int J Mol Sci 22:8462. https://doi.org/10.3390/ijms22168462
Del Pino-García R, Porrelli A, Rus-Fernández P, Segura-Carretero A, Curiel JA (2020) Identification, purification and characterization of a novel glycosidase (BgLm1) from Leuconostoc mesenteroides. LWT 122:108829. https://doi.org/10.1016/j.lwt.2019.108829
Escamilla-Lozano Y, Guzmán-Rodríguez F, Alatorre-Santamaría S, García-Garibay M, Gómez-Ruiz L, Rodríguez-Serrano G, Cruz-Guerrero A (2019) Synthesis of fucosyl-oligosaccharides using α-L-fucosidase from Lactobacillus rhamnosus GG. Molecules 24:2402. https://doi.org/10.3390/molecules24132402
Fernández-Lorente G, Bolívar JM, Rocha-Martin J, Curiel JA, Muñoz R, de Las RB, Carrascosa A, Guisan JM (2011) Synthesis of propyl gallate by transesterification of tannic acid in aqueous media catalysed by immobilised derivatives of tannase from Lactobacillus plantarum. Food Chem 128:214–217. https://doi.org/10.1016/j.foodchem.2011.02.057
Landete JM, Curiel JA, Rodríguez H, de las Rivas B, Muñoz R, (2014) Aryl glycosidases from Lactobacillus plantarum increase antioxidant activity of phenolic compounds. J Funct Foods 7:322–329. https://doi.org/10.1016/j.jff.2014.01.028
Li T, Li M, Hou L, Guo Y, Wang L, Sun G, Chen L (2018) Identification and characterization of a core fucosidase from the bacterium Elizabethkingia meningoseptica. J Biol Chem 293:1243–1258. https://doi.org/10.1074/jbc.M117.804252
Mateo C, Bolivar JM, Godoy CA, Rocha-Martin J, Pessela BC, Curiel JA, Muñoz R, Fernández-Lorente GJM, G, (2010) Improvement of enzyme properties with a two-step immobilizaton process on novel heterofunctional supports. Biomacromol 11(11):3112–3117. https://doi.org/10.1021/bm100916r
Rodríguez-Díaz J, Monedero V, Yebra MJ (2011) Utilization of natural fucosylated oligosaccharides by three novel α-l-fucosidases from a probiotic Lactobacillus casei strain. Appl Environ Microbiol 77:703–705. https://doi.org/10.1128/AEM.01906-10
Rodríguez-Díaz J, Rubio-del-Campo A, Yebra MJ (2012) Lactobacillus casei ferments the N-acetylglucosamine moiety of fucosyl-alpha-1,3-N-acetylglucosamine and excretes L-fucose. Appl Environ Microbiol 78:4613–4619. https://doi.org/10.1128/AEM.00474-12
Rodríguez-Díaz J, Carbajo RJ, Pineda-Lucena A, Monedero V, Yebra MJ (2013) Synthesis of fucosyl-N-acetylglucosamine disaccharides by transfucosylation using α-L-fucosidases from Lactobacillus casei. Appl Environ Microbiol 79:3847–3850. https://doi.org/10.1128/AEM.00229-13
Rozès N, Peres C (1998) Effects of phenolic compounds on the growth and the fatty acid composition of Lactobacillus plantarum. Appl Microbiol Biotechnol 49:108–111. https://doi.org/10.1007/s002530051145
Salli K, Hirvonen J, Siitonen J, Ahonen I, Anglenius H, Maukonen J (2021) Selective utilization of the human milk oligosaccharides 2′-fucosyllactose, 3-fucosyllactose, and difucosyllactose by various probiotic and pathogenic bacteria. J Agric Food Chem 69:170–182. https://doi.org/10.1021/acs.jafc.0c06041
Shi R, Ma J, Yan Q, Yang S, Fan Z, Jiang Z (2020) Biochemical characterization of a novel α-L-fucosidase from Pedobacter sp. and its application in synthesis of 3′-fucosyllactose and 2′-fucosyllactose. Appl Microbiol Biotechnol 104:5813–5826. https://doi.org/10.1007/s00253-020-10630-y
Verkhnyatskaya SA, Kong C, Klostermann CE, Schols HA, De Vos P, Walvoort MT (2021) Digestion, fermentation, and pathogen anti-adhesive properties of the hMO-mimic di-fucosyl-β-cyclodextrin. Food Funct 12:5018–5026. https://doi.org/10.1039/D1FO00830G
Zhang X, Mushajiang S, Luo B, Tian F, Ni Y, Yan W (2020) The composition and concordance of lactobacillus populations of infant gut and the corresponding breast-milk and maternal gut. Front Microbiol 2985. https://doi.org/10.3389/fmicb.2020.5
Zhou W, Jiang H, Wang L, Liang X, Mao X (2021) Biotechnological production of 2′-fucosyllactose: a prevalent fucosylated human milk oligosaccharide. ACS Synth Biol 10(3):447–458. https://doi.org/10.1021/acssynbio.0c00645
Zhu Y, Wan L, Li W, Ni D, Zhang W, Yan X, Mu W (2022). Recent advances on 2′-fucosyllactose: physiological properties, applications, and production approaches. Crit Rev Food Sci Nutr 62(8):2083–2092. 97911. https://doi.org/10.1080/10408398.2020.1850413
Zúñiga M, Monedero V, Yebra MJ (2018) Utilization of host-derived glycans by intestinal Lactobacillus and Bifidobacterium species. Front Microbiol 1917. https://doi.org/10.3389/fmicb.2018.01917
Acknowledgements
The authors are very grateful to the Analysis Service Unit (USTA-ICTAN) for the chromatography analysis.
Funding
This research was funded by the Spanish Ministry of Science and Innovation grants PID2020-119630RB-I00 and RYC2019-026368-I.
Author information
Authors and Affiliations
Contributions
JAC contributed to the study conception and design. Material preparation, data collection, and analysis were performed by AP, SL, EdV, and LB. The first draft of the manuscript was written by JAC, and JML commented on previous versions of the manuscript. All the authors read and approved the final manuscript.
Corresponding author
Ethics declarations
Ethical approval
This article does not contain any studies with human participants or animals performed by any of the authors.
Competing interests
The authors declare no competing interests.
Additional information
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
Below is the link to the electronic supplementary material.
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Curiel, J.A., Peirotén, Á., Langa, S. et al. Characterization and stabilization of the α-L-fucosidase set from Lacticaseibacillus rhamnosus INIA P603. Appl Microbiol Biotechnol 106, 8067–8077 (2022). https://doi.org/10.1007/s00253-022-12262-w
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-022-12262-w