CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function
ABSTRACT Invasive aspergillosis is one of the most serious clinical invasive fungal infections, resulting in a high case fatality rate among immunocompromised patients. The disease is caused by saprophytic molds in the genus Aspergillus, including Aspergillus fumigatus, the most significant pathogen...
| Main Authors: | , , , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
American Society for Microbiology
2023-08-01
|
| Series: | mBio |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mbio.00414-23 |
| _version_ | 1797730235984117760 |
|---|---|
| author | Xu Han Cecilia D'Angelo Ainara Otamendi Javier O. Cifuente Elisa de Astigarraga Borja Ochoa-Lizarralde Martin Grininger Francoise H. Routier Marcelo E. Guerin Jana Fuehring Oier Etxebeste Sean R. Connell |
| author_facet | Xu Han Cecilia D'Angelo Ainara Otamendi Javier O. Cifuente Elisa de Astigarraga Borja Ochoa-Lizarralde Martin Grininger Francoise H. Routier Marcelo E. Guerin Jana Fuehring Oier Etxebeste Sean R. Connell |
| author_sort | Xu Han |
| collection | DOAJ |
| description | ABSTRACT Invasive aspergillosis is one of the most serious clinical invasive fungal infections, resulting in a high case fatality rate among immunocompromised patients. The disease is caused by saprophytic molds in the genus Aspergillus, including Aspergillus fumigatus, the most significant pathogenic species. The fungal cell wall, an essential structure mainly composed of glucan, chitin, galactomannan, and galactosaminogalactan, represents an important target for the development of antifungal drugs. UDP (uridine diphosphate)-glucose pyrophosphorylase (UGP) is a central enzyme in the metabolism of carbohydrates that catalyzes the biosynthesis of UDP-glucose, a key precursor of fungal cell wall polysaccharides. Here, we demonstrate that the function of UGP is vital for Aspergillus nidulans (AnUGP). To understand the molecular basis of AnUGP function, we describe a cryoEM structure (global resolution of 3.5 Å for the locally refined subunit and 4 Å for the octameric complex) of a native AnUGP. The structure reveals an octameric architecture with each subunit comprising an N-terminal α-helical domain, a central catalytic glycosyltransferase A-like (GT-A-like) domain, and a C-terminal (CT) left-handed β-helix oligomerization domain. AnUGP displays unprecedented conformational variability between the CT oligomerization domain and the central GT-A-like catalytic domain. In combination with activity measurements and bioinformatics analysis, we unveil the molecular mechanism of substrate recognition and specificity for AnUGP. Altogether, our study not only contributes to understanding the molecular mechanism of catalysis/regulation of an important class of enzymes but also provides the genetic, biochemical, and structural groundwork for the future exploitation of UGP as a potential antifungal target. IMPORTANCE Fungi cause diverse diseases in humans, ranging from allergic syndromes to life-threatening invasive diseases, together affecting more than a billion people worldwide. Increasing drug resistance in Aspergillus species represents an emerging global health threat, making the design of antifungals with novel mechanisms of action a worldwide priority. The cryoEM structure of UDP (uridine diphosphate)-glucose pyrophosphorylase (UGP) from the filamentous fungus Aspergillus nidulans reveals an octameric architecture displaying unprecedented conformational variability between the C-terminal oligomerization domain and the central glycosyltransferase A-like catalytic domain in the individual protomers. While the active site and oligomerization interfaces are more highly conserved, these dynamic interfaces include motifs restricted to specific clades of filamentous fungi. Functional study of these motifs could lead to the definition of new targets for antifungals inhibiting UGP activity and, thus, the architecture of the cell wall of filamentous fungal pathogens. |
| first_indexed | 2024-03-12T11:41:19Z |
| format | Article |
| id | doaj.art-232e426a0a684ce694dd55e5be755b6b |
| institution | Directory Open Access Journal |
| issn | 2150-7511 |
| language | English |
| last_indexed | 2024-03-12T11:41:19Z |
| publishDate | 2023-08-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mBio |
| spelling | doaj.art-232e426a0a684ce694dd55e5be755b6b2023-08-31T15:04:20ZengAmerican Society for MicrobiologymBio2150-75112023-08-0114410.1128/mbio.00414-23CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and functionXu Han0Cecilia D'Angelo1Ainara Otamendi2Javier O. Cifuente3Elisa de Astigarraga4Borja Ochoa-Lizarralde5Martin Grininger6Francoise H. Routier7Marcelo E. Guerin8Jana Fuehring9Oier Etxebeste10Sean R. Connell11Structural Biology of Cellular Machines Laboratory, Biocruces Bizkaia Health Research Institute, Cruces University Hospital , Barakaldo, Bizkaia, SpainCenter for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA) , Derio, SpainLaboratory of Biology, Department of Applied Chemistry, Faculty of Chemistry, University of the Basque Country, UPV/EHU , San Sebastian, SpainCenter for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA) , Derio, SpainStructural Biology of Cellular Machines Laboratory, Biocruces Bizkaia Health Research Institute, Cruces University Hospital , Barakaldo, Bizkaia, SpainStructural Biology of Cellular Machines Laboratory, Biocruces Bizkaia Health Research Institute, Cruces University Hospital , Barakaldo, Bizkaia, SpainInstitute of Organic Chemistry and Chemical Biology, Buchmann Institute for Molecular Life Sciences, Goethe University Frankfurt , Frankfurt am Main, GermanyInstitute for Clinical Biochemistry, Hannover Medical School , Hannover, GermanyCenter for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA) , Derio, SpainInstitute for Clinical Biochemistry, Hannover Medical School , Hannover, GermanyLaboratory of Biology, Department of Applied Chemistry, Faculty of Chemistry, University of the Basque Country, UPV/EHU , San Sebastian, SpainStructural Biology of Cellular Machines Laboratory, Biocruces Bizkaia Health Research Institute, Cruces University Hospital , Barakaldo, Bizkaia, SpainABSTRACT Invasive aspergillosis is one of the most serious clinical invasive fungal infections, resulting in a high case fatality rate among immunocompromised patients. The disease is caused by saprophytic molds in the genus Aspergillus, including Aspergillus fumigatus, the most significant pathogenic species. The fungal cell wall, an essential structure mainly composed of glucan, chitin, galactomannan, and galactosaminogalactan, represents an important target for the development of antifungal drugs. UDP (uridine diphosphate)-glucose pyrophosphorylase (UGP) is a central enzyme in the metabolism of carbohydrates that catalyzes the biosynthesis of UDP-glucose, a key precursor of fungal cell wall polysaccharides. Here, we demonstrate that the function of UGP is vital for Aspergillus nidulans (AnUGP). To understand the molecular basis of AnUGP function, we describe a cryoEM structure (global resolution of 3.5 Å for the locally refined subunit and 4 Å for the octameric complex) of a native AnUGP. The structure reveals an octameric architecture with each subunit comprising an N-terminal α-helical domain, a central catalytic glycosyltransferase A-like (GT-A-like) domain, and a C-terminal (CT) left-handed β-helix oligomerization domain. AnUGP displays unprecedented conformational variability between the CT oligomerization domain and the central GT-A-like catalytic domain. In combination with activity measurements and bioinformatics analysis, we unveil the molecular mechanism of substrate recognition and specificity for AnUGP. Altogether, our study not only contributes to understanding the molecular mechanism of catalysis/regulation of an important class of enzymes but also provides the genetic, biochemical, and structural groundwork for the future exploitation of UGP as a potential antifungal target. IMPORTANCE Fungi cause diverse diseases in humans, ranging from allergic syndromes to life-threatening invasive diseases, together affecting more than a billion people worldwide. Increasing drug resistance in Aspergillus species represents an emerging global health threat, making the design of antifungals with novel mechanisms of action a worldwide priority. The cryoEM structure of UDP (uridine diphosphate)-glucose pyrophosphorylase (UGP) from the filamentous fungus Aspergillus nidulans reveals an octameric architecture displaying unprecedented conformational variability between the C-terminal oligomerization domain and the central glycosyltransferase A-like catalytic domain in the individual protomers. While the active site and oligomerization interfaces are more highly conserved, these dynamic interfaces include motifs restricted to specific clades of filamentous fungi. Functional study of these motifs could lead to the definition of new targets for antifungals inhibiting UGP activity and, thus, the architecture of the cell wall of filamentous fungal pathogens.https://journals.asm.org/doi/10.1128/mbio.00414-23aspergillosiscell wall biosynthesisUDP-glucose pyrophosphorylasesenzyme mechanismenzyme specificitycryoEM |
| spellingShingle | Xu Han Cecilia D'Angelo Ainara Otamendi Javier O. Cifuente Elisa de Astigarraga Borja Ochoa-Lizarralde Martin Grininger Francoise H. Routier Marcelo E. Guerin Jana Fuehring Oier Etxebeste Sean R. Connell CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function mBio aspergillosis cell wall biosynthesis UDP-glucose pyrophosphorylases enzyme mechanism enzyme specificity cryoEM |
| title | CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function |
| title_full | CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function |
| title_fullStr | CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function |
| title_full_unstemmed | CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function |
| title_short | CryoEM analysis of the essential native UDP-glucose pyrophosphorylase from Aspergillus nidulans reveals key conformations for activity regulation and function |
| title_sort | cryoem analysis of the essential native udp glucose pyrophosphorylase from aspergillus nidulans reveals key conformations for activity regulation and function |
| topic | aspergillosis cell wall biosynthesis UDP-glucose pyrophosphorylases enzyme mechanism enzyme specificity cryoEM |
| url | https://journals.asm.org/doi/10.1128/mbio.00414-23 |
| work_keys_str_mv | AT xuhan cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT ceciliadangelo cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT ainaraotamendi cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT javierocifuente cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT elisadeastigarraga cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT borjaochoalizarralde cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT martingrininger cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT francoisehroutier cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT marceloeguerin cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT janafuehring cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT oieretxebeste cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction AT seanrconnell cryoemanalysisoftheessentialnativeudpglucosepyrophosphorylasefromaspergillusnidulansrevealskeyconformationsforactivityregulationandfunction |