Bifunctional enhancement of a beta-glucanase-xylanase fusion enzyme by optimization of peptide linkers.
The flexible peptides (GGGGS)n (n < or = 3), the alpha-helical peptides (EAAAK)n (n < or = 3) and two other peptides were used as linkers to construct bifunctional fusions of beta-glucanase (Glu) and xylanase (Xyl) for improved catalytic efficiencies of both moieties. Eight Glu-Xyl fusion enzymes constructed with different linkers were all expressed as the proteins of ca. 46 kDa in Escherichia coli BL21 and displayed the activities of both beta-glucanase and xylanase. Compared to all the characterized fusions with the parental enzymes, the catalytic efficiencies of the Glu and Xyl moieties were equivalent to 304-426% and 82-143% of the parental ones, respectively. The peptide linker (GGGGS)(2) resulted in the best fusion, whose catalytic efficiency had a net increase of 326% for the Glu and of 43% for the Xyl. The two moieties of a fusion with the linker (EAAAK)(3) also showed net increases of 262 and 31% in catalytic efficiency. Our results highlight, for the first time, the enhanced bifunctional activities of the Glu-Xyl fusion enzyme by optimizing the peptide linkers to separate the two moieties at a reasonable distance for beneficial interaction.
Lu
Ping
P
Institute of Microbiology, College of Life Sciences, Zhejiang University, Hangzhou, Zhejiang, PR China.
Feng
Ming-Guang
MG
D001407:Bacillus / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013696:Temperature; D014990:Xylans / Q000378:metabolism; D047071:beta-Glucans / Q000378:metabolism
D001426:Bacterial Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; D014990:Xylans; D047071:beta-Glucans; D006026:Glycoside Hydrolases