title: Effect of Trichoderma reesei proteinases on the affinity of an inorganic-binding peptide.
abstract: An inorganic-binding peptide sequence with high affinity to silica-containing materials was fused to a glycoside hydrolase GH26 mannanase, ManA, from the extremely thermophilic bacterium Dictyoglomus thermophilum. The resulting recombinant enzyme produced in Escherichia coli, ManA-Linker, displayed high binding affinity towards synthetic zeolite while retaining its catalytic activity at 80 °C. ManA-Linker was able to bind to the zeolite at different pH levels, indicating a true pH-independent binding. However, complete degradation of the peptide linker was observed when the recombinant ManA-Linker was exposed to the supernatant from the filamentous fungus Trichoderma reesei. This degradation was caused by extracellular proteinases produced by T. reesei during its growth phase. Several derivatives of ManA-Linker were designed and expressed in E. coli. All the derivatives carrying a single sequence of the linker were still susceptible to T. reesei proteinase degradation. Complete substitution of the linker sequence by (GGGGS)16 resulted in a proteinase-resistant ManA derivative, ManA-Linker-(GGGGS)16, which was able to bind to zeolite in a pH-dependent manner.
authors: Department of Chemistry and Biomolecular Sciences, Faculty of Science, Macquarie University, North Ryde, Sydney, NSW 2109, Australia.
authors: Andrew
authors: A
authors: Care
authors: Helena
authors: H
authors: Nevalainen
authors: Peter L
authors: PL
authors: Bergquist
authors: Anwar
authors: A
authors: Sunna
mesh_terms: D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010447:Peptide Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014242:Trichoderma / Q000201:enzymology* / Q000235:genetics; D017641:Zeolites / Q000737:chemistry / Q000378:metabolism
chemical_list: D005656:Fungal Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; D017641:Zeolites; D010447:Peptide Hydrolases