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Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose

Lactococcus lactis strains are used as starter cultures in the production of fermented dairy and vegetable foods, but the species also occurs in other niches such as plant material. Lactococcus lactis subsp. lactis G50 (G50) is a plant-derived strain and potential candidate probiotics. Western blott...

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Autores principales: Tsuchiya, Wataru, Fujimoto, Zui, Inagaki, Noritoshi, Nakagawa, Hiroyuki, Tanaka, Miwa, Kimoto-Nira, Hiromi, Yamazaki, Toshimasa, Suzuki, Chise
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9815576/
https://www.ncbi.nlm.nih.gov/pubmed/36602978
http://dx.doi.org/10.1371/journal.pone.0273955
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author Tsuchiya, Wataru
Fujimoto, Zui
Inagaki, Noritoshi
Nakagawa, Hiroyuki
Tanaka, Miwa
Kimoto-Nira, Hiromi
Yamazaki, Toshimasa
Suzuki, Chise
author_facet Tsuchiya, Wataru
Fujimoto, Zui
Inagaki, Noritoshi
Nakagawa, Hiroyuki
Tanaka, Miwa
Kimoto-Nira, Hiromi
Yamazaki, Toshimasa
Suzuki, Chise
author_sort Tsuchiya, Wataru
collection PubMed
description Lactococcus lactis strains are used as starter cultures in the production of fermented dairy and vegetable foods, but the species also occurs in other niches such as plant material. Lactococcus lactis subsp. lactis G50 (G50) is a plant-derived strain and potential candidate probiotics. Western blotting of cell-wall proteins using antibodies generated against whole G50 cells detected a 120-kDa protein. MALDI-TOF MS analysis identified it as YwfG, a Leu-Pro-any-Thr-Gly cell-wall-anchor-domain–containing protein. Based on a predicted domain structure, a recombinant YwfG variant covering the N-terminal half (aa 28–511) of YwfG (YwfG(28−511)) was crystallized and the crystal structure was determined. The structure consisted of an L-type lectin domain, a mucin-binding protein domain, and a mucus-binding protein repeat. Recombinant YwfG variants containing combinations of these domains (YwfG(28–270), YwfG(28–336), YwfG(28−511), MubR4) were prepared and their interactions with monosaccharides were examined by isothermal titration calorimetry; the only interaction observed was between YwfG(28–270), which contained the L-type lectin domain, and d-mannose. Among four mannobioses, α-1,2-mannobiose had the highest affinity for YwfG(28–270) (dissociation constant = 34 μM). YwfG(28–270) also interacted with yeast mannoproteins and yeast mannan. Soaking of the crystals of YwfG(28–511) with mannose or α-1,2-mannobiose revealed that both sugars bound to the L-type lectin domain in a similar manner, although the presence of the mucin-binding protein domain and the mucus-binding protein repeat within the recombinant protein inhibited the interaction between the L-type lectin domain and mannose residues. Three of the YwfG variants (except MubR4) induced aggregation of yeast cells. Strain G50 also induced aggregation of yeast cells, which was abolished by deletion of ywfG from G50, suggesting that surface YwfG contributes to the interaction with yeast cells. These findings provide new structural and functional insights into the interaction between L. lactis and its ecological niche via binding of the cell-surface protein YwfG with mannose.
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spelling pubmed-98155762023-01-06 Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose Tsuchiya, Wataru Fujimoto, Zui Inagaki, Noritoshi Nakagawa, Hiroyuki Tanaka, Miwa Kimoto-Nira, Hiromi Yamazaki, Toshimasa Suzuki, Chise PLoS One Research Article Lactococcus lactis strains are used as starter cultures in the production of fermented dairy and vegetable foods, but the species also occurs in other niches such as plant material. Lactococcus lactis subsp. lactis G50 (G50) is a plant-derived strain and potential candidate probiotics. Western blotting of cell-wall proteins using antibodies generated against whole G50 cells detected a 120-kDa protein. MALDI-TOF MS analysis identified it as YwfG, a Leu-Pro-any-Thr-Gly cell-wall-anchor-domain–containing protein. Based on a predicted domain structure, a recombinant YwfG variant covering the N-terminal half (aa 28–511) of YwfG (YwfG(28−511)) was crystallized and the crystal structure was determined. The structure consisted of an L-type lectin domain, a mucin-binding protein domain, and a mucus-binding protein repeat. Recombinant YwfG variants containing combinations of these domains (YwfG(28–270), YwfG(28–336), YwfG(28−511), MubR4) were prepared and their interactions with monosaccharides were examined by isothermal titration calorimetry; the only interaction observed was between YwfG(28–270), which contained the L-type lectin domain, and d-mannose. Among four mannobioses, α-1,2-mannobiose had the highest affinity for YwfG(28–270) (dissociation constant = 34 μM). YwfG(28–270) also interacted with yeast mannoproteins and yeast mannan. Soaking of the crystals of YwfG(28–511) with mannose or α-1,2-mannobiose revealed that both sugars bound to the L-type lectin domain in a similar manner, although the presence of the mucin-binding protein domain and the mucus-binding protein repeat within the recombinant protein inhibited the interaction between the L-type lectin domain and mannose residues. Three of the YwfG variants (except MubR4) induced aggregation of yeast cells. Strain G50 also induced aggregation of yeast cells, which was abolished by deletion of ywfG from G50, suggesting that surface YwfG contributes to the interaction with yeast cells. These findings provide new structural and functional insights into the interaction between L. lactis and its ecological niche via binding of the cell-surface protein YwfG with mannose. Public Library of Science 2023-01-05 /pmc/articles/PMC9815576/ /pubmed/36602978 http://dx.doi.org/10.1371/journal.pone.0273955 Text en © 2023 Tsuchiya et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Tsuchiya, Wataru
Fujimoto, Zui
Inagaki, Noritoshi
Nakagawa, Hiroyuki
Tanaka, Miwa
Kimoto-Nira, Hiromi
Yamazaki, Toshimasa
Suzuki, Chise
Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose
title Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose
title_full Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose
title_fullStr Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose
title_full_unstemmed Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose
title_short Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose
title_sort cell-surface protein ywfg of lactococcus lactis binds to α-1,2-linked mannose
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9815576/
https://www.ncbi.nlm.nih.gov/pubmed/36602978
http://dx.doi.org/10.1371/journal.pone.0273955
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