Heterogeneity of homologously expressed Hypocrea jecorina (Trichoderma reesei) Cel7B catalytic module

• The catalytic module of Hypocrea jecorina (previously Trichoderma reesei) Cel7B was homologously expressed by transformation of strain QM9414. Post-translational modifications in purified Cel7B preparations were analysed by enzymatic digestions, high performance chromatography, mass spectrometry and site-directed mutagenesis. Of the five potential sites found in the wild-type enzyme, only Asn56 and Asn182 were found to be N-glycosylated. GlcNAc(2)Man(5) was identified as the predominant N-glycan, although lesser amounts of GlcNAc(2)Man(7) and glycans carrying a mannophosphodiester bond were also detected. Repartition of neutral and charged glycan structures over the two glycosylation sites mainly accounts for the observed microheterogeneity of the protein. However, partial deamidation of Asn259 and a partially occupied O-glycosylation site give rise to further complexity in enzyme preparations.

Ämnesord

NATURVETENSKAP  -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)

NATURVETENSKAP  -- Biologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences (hsv//eng)

Nyckelord

protein glycosylation

O-glycan

N-glycan

Trichoderma reesei

cellulase

cellobiohydrolase-i cel7a

aqueous 2-phase systems

endoglucanase-i

asparagine residues

crystal-structure

n-glycosylation

sequence

protein

identification

deamidation

N-glycan

protein glycosylation

O-glycan

Tricho-derma reesei

cellulase.

Publikations- och innehållstyp

ref (ämneskategori)

art (ämneskategori)