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Metabolic Engineering of the Morphology of Aspergillus oryzae by Altering Chitin Synthesis

机译:改变几丁质合成的米曲霉形态的代谢工程。

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摘要

Morphology and α-amylase production during submerged cultivation were examined in a wild-type strain (A1560) and in strains of Aspergillus oryzae in which chitin synthase B (chsB) and chitin synthesis myosin A (csmA) have been disrupted (ChsB/G and CM101). In a flowthrough cell, the growth of submerged hyphal elements was studied online, making it possible to examine the growth kinetics of the three strains. The average tip extension rates of the CM101 and ChsB/G strains were 25 and 88% lower, respectively, than that of the wild type. The branching intensity in the CM101 strain was 25% lower than that in the wild type, whereas that in the ChsB/G strain was 188% higher. During batch cultivation, inseparable clumps were formed in the wild-type strain, while no or fewer large inseparable clumps existed in the cultivations of the ChsB/G and CM101 strains. The α-amylase productivity was not significantly different in the three strains. A strain in which the transcription of chsB could be controlled by the nitrogen source-regulated promoter niiA (NiiA1) was examined during chemostat cultivation, and it was found that the branching intensity could be regulated by regulating the promoter, signifying an important role for chsB in branching. However, the pattern of branching responded very slowly to the change in transcription, and increased branching did not affect α-amylase productivity. α-Amylase residing in the cell wall was stained by immunofluorescence, and the relationship between tip number and enzyme secretion is discussed.
机译:在野生型菌株(A1560)和米曲霉菌株中检测了淹没培养期间的形态和α-淀粉酶产生,其中几丁质合酶B(chsB)和几丁质合成肌球蛋白A(csmA)受到破坏(ChsB / G和CM101)。在流通池中,在线研究了淹没菌丝元素的生长,从而有可能检查这三种菌株的生长动力学。 CM101和ChsB / G菌株的平均末端延伸率分别比野生型低25和88%。 CM101菌株的分支强度比野生型低25%,而ChsB / G菌株的分支强度高188%。在分批培养过程中,野生型菌株中形成了不可分离的团块,而ChsB / G和CM101菌株的培养中不存在或几乎没有大的不可分离的团块。在三种菌株中,α-淀粉酶生产率没有显着差异。在化学恒温恒湿培养过程中检查了一种可以通过氮源调控的启动子niiA(NiiA1)控制chsB转录的菌株,发现可以通过调节启动子来调节分支强度,这对于chsB具有重要作用。在分支。但是,分支模式对转录变化的响应非常慢,并且增加的分支不会影响α-淀粉酶的生产率。通过免疫荧光法对细胞壁中的α-淀粉酶进行染色,探讨了尖端数目与酶分泌之间的关系。

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