Increased enzyme production under liquid culture conditions in the industrial fungus <i>Aspergillus oryzae</i> by disruption of the genes encoding cell wall α-1,3-glucan synthase
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Under liquid culture conditions, the hyphae of filamentous fungi aggregate to form pellets, which reduces cell density and fermentation productivity. Previously, we found that loss of α-1,3-glucan in the cell wall of the fungus <i>Aspergillus nidulans</i> increased hyphal dispersion. Therefore, here we constructed a mutant of the industrial fungus <i>A. oryzae</i> in which the three genes encoding α-1,3-glucan synthase were disrupted (tripleΔ). Although the hyphae of the tripleΔ mutant were not fully dispersed, the mutant strain did form smaller pellets than the wild-type strain. We next examined enzyme productivity under liquid culture conditions by transforming the cutinase-encoding gene <i>cutL1</i> into <i>A. oryzae</i> wild-type and the tripleΔ mutant (i.e. wild-type-cutL1, tripleΔ-cutL1). <i>A. oryzae</i> tripleΔ-cutL1 formed smaller hyphal pellets and showed both greater biomass and increased CutL1 productivity compared with wild-type-cutL1, which might be attributable to a decrease in the number of tripleΔ-cutL1 cells under anaerobic conditions. Summary of the growth characteristic and enzyme productivity in <i>A. oryzae ags</i> tripleΔ strain.
在液体培养条件下，丝状真菌的菌丝会聚集形成菌丝球（pellets），这一现象会降低细胞密度与发酵生产效率。本课题组此前发现，构巢曲霉（*Aspergillus nidulans*）细胞壁中α-1,3-葡聚糖（α-1,3-glucan）的缺失可增强菌丝的分散性。据此，本研究构建了工业用真菌米曲霉（*Aspergillus oryzae*，简称A. oryzae）的三基因敲除突变体，该突变体中3个编码α-1,3-葡聚糖合酶（α-1,3-glucan synthase）的基因被成功敲除（以下简称tripleΔ突变体）。尽管tripleΔ突变体的菌丝并未完全分散，但该突变菌株所形成的菌丝球尺寸显著小于野生型菌株。随后，本研究将编码角质酶（cutinase）的*cutL1*基因分别转化至米曲霉野生型菌株与tripleΔ突变体中，获得wild-type-cutL1与tripleΔ-cutL1工程菌株，并以此检测液体培养条件下的酶生产效率。与wild-type-cutL1菌株相比，米曲霉tripleΔ-cutL1菌株不仅形成了尺寸更小的菌丝球，同时具备更高的生物量与更强的CutL1酶生产能力，该现象可能与厌氧条件下tripleΔ-cutL1菌株的细胞数量减少有关。米曲霉ags tripleΔ菌株的生长特性与酶生产效率总结。
