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Cloning and characterization of a putative β-glucosidase (NfBGL595) from Neosartorya fischeri

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Whole genome sequence of Neosartorya fischeri NRRL181 revealed four putative GH1 β-glucosidases (BGLs). One BGL, NfBGL595 was successfully expressed and characterized. DNA sequence analysis revealed an open reading frame of 1590 bp, encoding a polypeptide of 529 amino acid residues. The gene was cloned in pET28a and overexpressed in Escherichia coli. The purified recombinant BGL showed high levels of catalytic activity, with V max of 1693 U mg-protein -1 and a K m of 2.8 mM for p-nitrophenyl-β-d-glucopyranoside (pNPG). The optimal temperature and pH for enzyme activity were 40 °C and 6.0, respectively. The enzyme exhibited broad substrate specificity towards aryl glycosides including pNP-mannose, pNP-galactose, pNP-xylose, and pNP-cellobioside. A homology model of NfBGL595 was constructed based on the X-ray crystal structure of Trichoderma reesei BGL2. Molecular dynamics simulation studies of the enzyme with the pNPG and cellobiose, shed light on the substrate specificity of N. fischeri BGL595 only towards aryl glycoside.

OriginalsprogEngelsk
TidsskriftProcess Biochemistry
Vol/bind47
Udgave nummer1
Sider (fra-til)99-105
Antal sider7
ISSN1359-5113
DOI
StatusUdgivet - 2012
Eksternt udgivetJa

ID: 203252031