The Gas family of proteins of Saccharomyces cerevisiae: characterization and evolutionary analysis.
The GAS multigene family of Saccharomyces cerevisiae is constituted by five genes (GAS1-GAS5), but GAS1 was the only one to have been characterized to date. Gas1 is a glycosylphosphatidylinositol-anchored protein predominantly localized in the plasma membrane and is also a representative of family GH72 of glycosidase/transglycosidases, a wide group of yeast and fungal enzymes involved in cell wall assembly. Gas1-Gas5 proteins share a common N-terminal domain but exhibit different C-terminal extensions, in which a domain named Cys-Box is located. This domain is similar to the carbohydrate binding module 43 and is present only in Gas1p and Gas2p. Here we report the expression in P. pastoris of soluble forms of Gas proteins. Gas1, 2, 4 and 5 proteins were secreted with a yield of about 30-40 mg/l of medium, whereas the yield for Gas3p was about three times lower. Gas proteins proved to be N-glycosylated. Purified Gas proteins were tested for enzymatic activity. Gas2, Gas4 and Gas5p showed a beta-(1,3)-glucanosyltransferase activity similar to Gas1p. A phylogenetic tree of the N-terminal regions of family GH72 members was constructed. Two subfamilies of N-terminal regions were distinguished: one subfamily, GH72(+), contains proteins that possess a Cys-box in the C-terminal region, whereas family GH72(-) comprises proteins that lack a Cys-box. On the basis of this net distinction, we speculate that the type of C-tail region imposed constraints to the evolution of the N-terminal portion.
Ragni
Enrico
E
Università degli Studi di Milano, Dipartimento di Scienze Biomolecolari e Biotecnologie, 20133 Milano, Italy.
Fontaine
Thierry
T
Gissi
Carmela
C
Latgè
Jean Paul
JP
Popolo
Laura
L
D002473:Cell Wall / Q000378:metabolism; D019143:Evolution, Molecular; D015966:Gene Expression Regulation, Fungal; D004693:Glucan Endo-1,3-beta-D-Glucosidase / Q000235:genetics / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D005810:Multigene Family; D010802:Phylogeny; D010843:Pichia / Q000201:enzymology / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism
C066709:GAS1 protein, S cerevisiae; D008562:Membrane Glycoproteins; D029701:Saccharomyces cerevisiae Proteins; C117939:1,3-beta-glucanosyltransferase; D004693:Glucan Endo-1,3-beta-D-Glucosidase