Mechanisms of redundancy and specificity of the Aspergillus fumigatus Crh transglycosylases

Wenxia Fang, Ana Belén Sanz, Sergio Galan Bartual, Bin Wang, Andrew Ferenbach, Vladimír Farkaš, Ramón Hurtado-Guerrero, Javier Arroyo (Lead / Corresponding author), Daan van Aalten (Lead / Corresponding author)

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Abstract

Fungal cell wall synthesis is achieved by a balance of glycosyltransferase, hydrolase and transglycosylase activities. Transglycosylases strengthen the cell wall by forming a rigid network of crosslinks through mechanisms that remain to be explored. Here we study the function of the Aspergillus fumigatus family of five Crh transglycosylases. Although crh genes are dispensable for cell viability, simultaneous deletion of all genes renders cells sensitive to cell wall interfering compounds. In vitro biochemical assays and localisation studies demonstrate that this family of enzymes functions redundantly as transglycosylases for both chitin-glucan and chitin-chitin cell wall crosslinks. To understand the molecular basis of this acceptor promiscuity, we solved the crystal structure of A. fumigatus Crh5 (AfCrh5) in complex with a chitooligosaccharide at the resolution of 2.8 Å, revealing an extensive elongated binding cleft for the donor (−4 to −1) substrate and a short acceptor (+1 to +2) binding site. Together with mutagenesis, the structure suggests a “hydrolysis product assisted” molecular mechanism favouring transglycosylation over hydrolysis.

Original languageEnglish
Article number1669
Pages (from-to)1-10
Number of pages10
JournalNature Communications
Volume10
Issue number1
DOIs
Publication statusPublished - 10 Apr 2019

Fingerprint

Aspergillus
Aspergillus fumigatus
redundancy
Cell Wall
Chitin
Redundancy
chitin
Cells
Hydrolysis
genes
hydrolysis
Glycosyltransferases
Genes
Glucans
Gene Deletion
Hydrolases
mutagenesis
deletion
Mutagenesis
Cell Survival

Keywords

  • Aspergillus fumigatus
  • cell wall
  • chitin
  • cross-link
  • enzymology
  • fungi
  • genetics
  • glucan
  • protein structure
  • transglycosylase

Cite this

Fang, Wenxia ; Sanz, Ana Belén ; Galan Bartual, Sergio ; Wang, Bin ; Ferenbach, Andrew ; Farkaš, Vladimír ; Hurtado-Guerrero, Ramón ; Arroyo, Javier ; van Aalten, Daan. / Mechanisms of redundancy and specificity of the Aspergillus fumigatus Crh transglycosylases. In: Nature Communications. 2019 ; Vol. 10, No. 1. pp. 1-10.
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Fang, W, Sanz, AB, Galan Bartual, S, Wang, B, Ferenbach, A, Farkaš, V, Hurtado-Guerrero, R, Arroyo, J & van Aalten, D 2019, 'Mechanisms of redundancy and specificity of the Aspergillus fumigatus Crh transglycosylases', Nature Communications, vol. 10, no. 1, 1669, pp. 1-10. https://doi.org/10.1038/s41467-019-09674-0

Mechanisms of redundancy and specificity of the Aspergillus fumigatus Crh transglycosylases. / Fang, Wenxia; Sanz, Ana Belén ; Galan Bartual, Sergio; Wang, Bin ; Ferenbach, Andrew; Farkaš, Vladimír ; Hurtado-Guerrero, Ramón; Arroyo, Javier (Lead / Corresponding author); van Aalten, Daan (Lead / Corresponding author).

In: Nature Communications, Vol. 10, No. 1, 1669, 10.04.2019, p. 1-10.

Research output: Contribution to journalArticle

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AU - Sanz, Ana Belén

AU - Galan Bartual, Sergio

AU - Wang, Bin

AU - Ferenbach, Andrew

AU - Farkaš, Vladimír

AU - Hurtado-Guerrero, Ramón

AU - Arroyo, Javier

AU - van Aalten, Daan

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N2 - Fungal cell wall synthesis is achieved by a balance of glycosyltransferase, hydrolase and transglycosylase activities. Transglycosylases strengthen the cell wall by forming a rigid network of crosslinks through mechanisms that remain to be explored. Here we study the function of the Aspergillus fumigatus family of five Crh transglycosylases. Although crh genes are dispensable for cell viability, simultaneous deletion of all genes renders cells sensitive to cell wall interfering compounds. In vitro biochemical assays and localisation studies demonstrate that this family of enzymes functions redundantly as transglycosylases for both chitin-glucan and chitin-chitin cell wall crosslinks. To understand the molecular basis of this acceptor promiscuity, we solved the crystal structure of A. fumigatus Crh5 (AfCrh5) in complex with a chitooligosaccharide at the resolution of 2.8 Å, revealing an extensive elongated binding cleft for the donor (−4 to −1) substrate and a short acceptor (+1 to +2) binding site. Together with mutagenesis, the structure suggests a “hydrolysis product assisted” molecular mechanism favouring transglycosylation over hydrolysis.

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