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首页> 外文期刊>Biotechnology Progress >High-Level Expression of Aspergillus niger P-Galactosidase in Ashbya gossypii
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High-Level Expression of Aspergillus niger P-Galactosidase in Ashbya gossypii

机译：黑曲霉P-半乳糖苷酶在棉铃虫中的高表达

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Ashbya gossypii has been recently considered as a host for the expression of recombinant proteins. The production levels achieved thus far were similar to those obtained with Sac-charomyces cerevisiae for the same proteins. Here, the β-galactosidase from Aspergillus niger was successfully expressed and secreted by A. gossypii from 2-im plasmids carrying the native signal sequence at higher levels than those secreted by S. cerevisiae laboratorial strains. Four different constitutive promoters were used to regulate the expression of β-galactosidase: A. gossypii AgTEF and AgGPD promoters, and S. cerevisiae ScADHl and ScPGKl promoters. The native AgTEF promoter drove the highest expression levels of recombinant β-galactosidase in A. gossypii, leading to 2-and 8-fold higher extracellular activity than the AgGPD promoter and the heterologous promoters, respectively. In similar production conditions, the levels of active β-galactosidase secreted by A. gossypii were up to 37 times higher than those secreted by recombinant S. cerevisiae and ~2.5 times higher than those previously reported for the β-galactosidase-high producing S. cerevisiae NCYC869-A3/pVKl .1. The substitution of glucose by glycerol in the production medium led to a 1.5-fold increase in the secretion of active β-galactosidase by A. gossypii. Recombinant β-galactosidase secreted by A. gossypii was extensively glycosylated, as are the native A. niger β-galactosidase and recombinant β-galactosidase produced by yeast. These results highlight the potential of A. gossypii as a recombinant protein producer and open new perspectives to further optimize recombinant protein secretion in this fungus.
机译：最近已将棉兰阿什比棉（Ashbya gossypii）用作表达重组蛋白的宿主。迄今为止，所获得的生产水平与酿酒酵母对相同蛋白质所获得的水平相似。在这里，来自黑曲霉的β-半乳糖苷酶已被棉孢曲霉成功地表达并分泌，其携带天然信号序列的2- im质粒比酿酒酵母实验室菌株分泌的水平更高。四种不同的组成型启动子用于调节β-半乳糖苷酶的表达：棉球菌AgTEF和AgGPD启动子，以及酿酒酵母ScADH1和ScPGK1启动子。天然的AgTEF启动子驱动棉球曲霉中重组β-半乳糖苷酶的最高表达水平，从而导致细胞外活性分别比AgGPD启动子和异源启动子高2和8倍。在相似的生产条件下，棉球菌分泌的活性β-半乳糖苷酶水平比重组酿酒酵母所分泌的活性β-半乳糖苷酶高37倍，比先前报道的高产β-半乳糖苷酶的S.高约2.5倍。酿酒酵母NCYC869-A3 / pVK1.1。生产培养基中的甘油替代葡萄糖导致棉球菌的活性β-半乳糖苷酶分泌增加了1.5倍。棉球菌分泌的重组β-半乳糖苷酶被广泛地糖基化，酵母产生的天然黑曲霉β-半乳糖苷酶和重组β-半乳糖苷酶也被广泛糖基化。这些结果突出了棉孢曲霉作为重组蛋白生产者的潜力，并为进一步优化该真菌中重组蛋白的分泌开辟了新的前景。

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