title: Purification, physico-chemical and kinetic properties of the deglycosylated Talaromyces thermophilus lipase.
abstract: The Talaromyces thermophilus strain produces only one form of lipase called TTLI. When the culture medium was concentrated and stored at 4°C during a few days, we noticed the appearance of a second short form of lipase named TTLII. This second form was purified to homogeneity using gel filtration and FPLC-Anion exchange chromatography. The NH(2)-terminal 24 amino acid residues were found to be identical to those of TTLI. The treatment of the TTLI with endoglycosidase H decreased its apparent molecular weight from 39 to 30kDa which corresponds to the molecular weight of TTLII. This difference was mostly attributed to the N-glycosylation of the enzyme. In fact, the glycan chain content and concavaline A-Sepharose affinity column confirmed that the TTLII was completely deglycosylated. Compared to TTLI, the TTLII activity was completely decreased over a broad range of temperature and pH. Furthermore, the deglycosylation of the enzyme reduced its specific activity by 50% toward different substrates; strongly suggest that the N-glycans are determinants for optimal catalytic activity and thermal stability of this enzyme. Covalent immobilization of the enzymes on supports suggests the involvement of the glycan moiety in enzyme-polymer interactions. In the case of TTLI the glycan moiety can constitute an extra site for the covalent linkage of the enzyme on the carrier.
authors: Laboratoire de Valorisation de la Biomasse et Production des Protéines chez les Eucaryotes, Centre de Biotechnologies de Sfax, BP «1177» 3018 Sfax, University of Sfax, Tunisia.
authors: Ines belhaj-ben
authors: Ib
authors: Romdhan
authors: Ahmed
authors: A
authors: Fendri
authors: Fakher
authors: F
authors: Frikha
authors: Ali
authors: A
authors: Gargouri
authors: Hafedh
authors: H
authors: Belghith
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D055162:Biocatalysis; D002241:Carbohydrates / Q000737:chemistry; D055598:Chemical Phenomena; D006031:Glycosylation; D007700:Kinetics; D008049:Lipase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D013379:Substrate Specificity; D032901:Talaromyces / Q000201:enzymology*
chemical_list: D002241:Carbohydrates; D008049:Lipase