title: N-glycosylated catalytic unit meets O-glycosylated propeptide: complex protein architecture in a fungal hexosaminidase.
abstract: beta-N-Acetylhexosaminidase from a filamentous fungus Aspergillus oryzae is a secreted enzyme known to be an important component of the binary chitinolytic system. Cloning of the hexA gene and sequencing of the enzyme revealed its unique preproprotein structure. While the enzyme's zincin-like and catalytic domain had significant similarities with members of the glycohydrolase 20 family, the propeptide was unique for the fungal enzyme. Detailed pulse-chase and inhibition studies revealed that propeptide was processed during the biosynthesis of the enzyme. Moreover, the presence of propeptide was necessary for enzyme activation, dimerization and secretion. The catalytic unit was N-glycosylated, and the propeptide was O-glycosylated, both in their C-terminal parts. Deglycosylation experiments revealed that the N-glycosylation increased the stability and solubility of the enzyme. In contrast, O-glycosylated propeptide was necessary to attain the full enzymic activity.
authors.affiliation: Institute of Microbiology, Academy of Sciences of the Czech Republic, Vídenská 1083, CZ-14220 Praha 4, Czech Republic. plihal@biomed.cas.cz
authors.forename: O
authors.initials: O
authors.lastname: Plíhal
authors.forename: J
authors.initials: J
authors.lastname: Sklenár
authors.forename: J
authors.initials: J
authors.lastname: Kmonícková
authors.forename: P
authors.initials: P
authors.lastname: Man
authors.forename: P
authors.initials: P
authors.lastname: Pompach
authors.forename: V
authors.initials: V
authors.lastname: Havlícek
authors.forename: V
authors.initials: V
authors.lastname: Kren
authors.forename: K
authors.initials: K
authors.lastname: Bezouska
mesh_terms: D001236:Aspergillus oryzae / Q000201:enzymology*; D020134:Catalytic Domain; D002686:Chitin / Q000737:chemistry; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D006031:Glycosylation; D054818:Hexosaminidase A; D006596:Hexosaminidases / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D010455:Peptides / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors; D001619:beta-N-Acetylhexosaminidases / Q000737:chemistry
chemical_list: D010455:Peptides; D002686:Chitin; D006596:Hexosaminidases; D054818:Hexosaminidase A; D001619:beta-N-Acetylhexosaminidases