The Gel/Gas/Phr family of fungal β(1,3)-glucanosyltransferases plays an important role in cell wall biogenesis by processing the main component β(1,3)-glucan. Two subfamilies are distinguished depending on the presence or absence of a C-terminal cysteine-rich domain, denoted "Cys-box." The N-terminal domain (NtD) contains the catalytic residues for transglycosidase activity and is separated from the Cys-box by a linker region. To obtain a better understanding of the structure and function of the Cys-box-containing subfamily, we identified the disulfide bonds in Gas2p from Saccharomyces cerevisiae by an improved mass spectrometric methodology. We mapped two separate intra-domain clusters of three and four disulfide bridges. One of the bonds in the first cluster connects a central Cys residue of the NtD with a single conserved Cys residue in the linker. Site-directed mutagenesis of the Cys residue in the linker resulted in an endoplasmic reticulum precursor that was not matured and underwent a gradual degradation. The relevant disulfide bond has a crucial role in folding as it may stabilize the NtD and facilitate its interaction with the C-terminal portion of a Gas protein. The four disulfide bonds in the Cys-box are arranged in a manner consistent with a partial structural resemblance with the plant X8 domain, an independent carbohydrate-binding module that possesses only three disulfide bonds. Deletion of the Cys-box in Gas2 or Gas1 proteins led to the formation of an NtD devoid of any enzymatic activity. The results suggest that the Cys-box is required for proper folding of the NtD and/or substrate binding.

Disulfide bond structure and domain organization of yeast beta(1,3)-glucanosyltransferases involved in cell wall biogenesis / L. Popolo, E. Ragni, C. Carotti, O. Palomares, R. Aardema, J.W. Back, H.L. Dekker, L.J. de Koning, L. de Jong, C.G. de Koster. - In: THE JOURNAL OF BIOLOGICAL CHEMISTRY. - ISSN 0021-9258. - 283:27(2008), pp. 18553-18565. [10.1074/jbc.M801562200]

Disulfide bond structure and domain organization of yeast beta(1,3)-glucanosyltransferases involved in cell wall biogenesis

L. Popolo
Primo
;
E. Ragni
Secondo
;
C. Carotti;
2008

Abstract

The Gel/Gas/Phr family of fungal β(1,3)-glucanosyltransferases plays an important role in cell wall biogenesis by processing the main component β(1,3)-glucan. Two subfamilies are distinguished depending on the presence or absence of a C-terminal cysteine-rich domain, denoted "Cys-box." The N-terminal domain (NtD) contains the catalytic residues for transglycosidase activity and is separated from the Cys-box by a linker region. To obtain a better understanding of the structure and function of the Cys-box-containing subfamily, we identified the disulfide bonds in Gas2p from Saccharomyces cerevisiae by an improved mass spectrometric methodology. We mapped two separate intra-domain clusters of three and four disulfide bridges. One of the bonds in the first cluster connects a central Cys residue of the NtD with a single conserved Cys residue in the linker. Site-directed mutagenesis of the Cys residue in the linker resulted in an endoplasmic reticulum precursor that was not matured and underwent a gradual degradation. The relevant disulfide bond has a crucial role in folding as it may stabilize the NtD and facilitate its interaction with the C-terminal portion of a Gas protein. The four disulfide bonds in the Cys-box are arranged in a manner consistent with a partial structural resemblance with the plant X8 domain, an independent carbohydrate-binding module that possesses only three disulfide bonds. Deletion of the Cys-box in Gas2 or Gas1 proteins led to the formation of an NtD devoid of any enzymatic activity. The results suggest that the Cys-box is required for proper folding of the NtD and/or substrate binding.
English
Settore BIO/11 - Biologia Molecolare
Articolo
Esperti anonimi
Pubblicazione scientifica
2008
American Society for Biochemistry and Molecular Biology
283
27
18553
18565
13
Pubblicato
Periodico con rilevanza internazionale
Aderisco
info:eu-repo/semantics/article
Disulfide bond structure and domain organization of yeast beta(1,3)-glucanosyltransferases involved in cell wall biogenesis / L. Popolo, E. Ragni, C. Carotti, O. Palomares, R. Aardema, J.W. Back, H.L. Dekker, L.J. de Koning, L. de Jong, C.G. de Koster. - In: THE JOURNAL OF BIOLOGICAL CHEMISTRY. - ISSN 0021-9258. - 283:27(2008), pp. 18553-18565. [10.1074/jbc.M801562200]
open
Prodotti della ricerca::01 - Articolo su periodico
10
262
Article (author)
Periodico con Impact Factor
L. Popolo, E. Ragni, C. Carotti, O. Palomares, R. Aardema, J.W. Back, H.L. Dekker, L.J. de Koning, L. de Jong, C.G. de Koster
File in questo prodotto:
File Dimensione Formato  
JBiolChem2008_2434_41588.pdf

accesso aperto

Tipologia: Publisher's version/PDF
Dimensione 733.78 kB
Formato Adobe PDF
733.78 kB Adobe PDF Visualizza/Apri
Pubblicazioni consigliate

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/41588
Citazioni
  • ???jsp.display-item.citation.pmc??? 10
  • Scopus 30
  • ???jsp.display-item.citation.isi??? 31
social impact