Mutagenesis of the glucoamylase signal peptide of Saccharomyces diastaticus and functional analysis in Saccharomyces cerevisiae |
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Authors: | Lee J W Kang D O Kim B Y Oh W K Mheen T I Pyun Y R Ahn J S |
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Institution: | Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejin University, Yusong, Taejon, South Korea. |
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Abstract: | To improve the efficiency of the glucoamylase signal peptide (GSP) of Saccharomyces diastaticus for the secretion of foreign proteins, hybrid plasmids containing one of four types of GSP mutant (m1, Pro(-18)-->Leu(-18); m2, Tyr(-13)-->Leu(-13); m3, Ser(-9)-->Leu(-9); m4, Asn(-5)-->Pro(-5)) were constructed and evaluated in Saccharomyces cerevisiae using Bacillus endo-1,4-beta-D-glucanase (CMCase) as a reporter gene. CMCase secretion by m1, m2 and m3 GSP mutants was increased, likely resulting from a higher probability of the modified GSP to assume an alpha-helical structure. Especially in the case of m3, the substitution of Leu for a polar residue, Ser(-9), in the hydrophobic region resulted in approximately a twofold increase in extracellular CMCase activity. In mutant 4, which disrupts the alpha-helix of GSP, CMCase was less efficiently secreted. |
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Keywords: | Mutagenesis Glucoamylase signal peptide Saccharomyces diastaticus Bacillus endo-1 4-β-D-glucanase Saccharomyces cerevisiae Secretion |
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