N-linked carbohydrate chains protect laccase III from proteolysis in Coriolus versicolor

Abstract

The structure and function of the N-linked carbohydrate chains in laccase 111, one of the ligninolytic glycoenzymes from the white-rot fungus Cuviulus versiculuv, have been partially characterized using endoglycosidases (Endo F and Endo H) and the N-asparagine-specific inhibitor, tunicamycin. In the presence of 10 pg tunicamycin ml-' laccase and proteinase activities in culture filtrates of C. vevsiculur were measured over 3 weeks. Laccase activity was slightly decreased by the addition of tunicamycin, whereas proteinase activity was increased. The N-linked carbohydrate chains were not necessary for laccase secretion and activity. Endo-glycosidase digestion showed that laccase I11 contained at least four N-linked carbohydrate chains, of which two were high-mannose type or hybrid type and two were complex type. Judging from the differences in the resistance of the native and the carbohydratedepleted laccase to proteolytic digestion and high temperature, the four N-linked carbohydrate chains have important protective functions against proteolytic attack and elevated temperature.

Stability Mechanisms of a Thermophilic Laccase Probed by Molecular Dynamics

“…To be discussed later, NaCl has a stabilizing effect showing up when glycosylation does not prevent anion diffusion into the protein. Absent this case, glycosylation consistently strengthens secondary structure hydrogen bonds, which is a possible explanation for the experimental findings that N-glycosylation confers thermostability to Trametes versicolor laccase isoform 3 [70] as also observed for other classes of proteins [71], [72], [73]. Our MD-simulations produce a significant (14 of compared 15 pairs of simulations averaged over 100 snapshots each) secondary-structure-preserving effect of glycosylation and relate it to an intriguing interplay between salt penetration and thermal disruption of secondary structure.…”

Fungal laccases – occurrence and properties

“…The glycosylation of fungal laccases is one of the biggest problems for the heterologous production of the enzyme, which is extremely difficult to overcome. It was proposed that in addition to the structural role, glycosylation can also participate in the protection of laccase from proteolytic degradation (Yoshitake et al , 1993).…”

Two laccase isoforms of the basidiomycete Cerrena unicolor VKMF‐3196. Induction, isolation and properties

“…An important factor determining the stability of laccase protein is the extent of its glycosilation. The carbohydrate moiety of laccases is often reffered to as a protective factor against proteolysis and thermoinactivation [42,43]. The larger amount of carbohydrates in the structure of LacC1 is a possible reason of its higher persistence to high temperature, as compared to LacC2.…”