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Reduction of Extracellular Proteases Increased Activity and Stability of Heterologous Protein in Aspergillus niger

Kamaruddin, Nurhaida and Storms, Reginald and Mahadi, Nor Muhammad and Md. Illias, Rosli and Abu Bakar, Farah Diba and Abdul Murad, Abdul Munir (2018) Reduction of Extracellular Proteases Increased Activity and Stability of Heterologous Protein in Aspergillus niger. Arabian Journal for Science and Engineering, 43 (7). pp. 3327-3338. ISSN 2193-567X

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Official URL: http://dx.doi.org/10.1007/s13369-017-2914-3

Abstract

The heterologous protein production in Aspergillus niger is often limited by the activity of the host extracellular proteases. To improve heterologous production, a transcription factor controlling expression of several extracellular protease-encoding genes, prtT, was deleted in A. niger PY11 and the mutant (AnΔ prtT) characterised. Extracellular proteolytic activity of AnΔ prtT was reduced as compared to the wild type, a result that was confirmed by RT-PCR analyses that showed reduced expression levels of several protease gene transcripts. To compare the efficiency of the mutant and parental (PY11) strains as hosts for heterologous protein production, the cutinase gene from Glomerella cingulate, under control of the glucoamylase A promoter, was integrated into each genome. The cutinase activity of PY11 and AnΔ prtT harbouring the G. cingulata cutinase gene was increased 20- and 36-fold higher, respectively, than the untransformed parental strains, suggesting that the ability of the mutant to produce heterologous protein is better than the wild type. Cutinase activity in culture filtrates prepared using both strains was stable at 4∘C for extended periods; however, during incubation at 25 ∘C the heterologous cutinase in culture filtrates prepared using AnΔ prtT retained 80% activity after a two-week incubation versus the less than 3% activity that was retained in culture filtrates prepared using PY11. This indicates that the reduction of extracellular proteases greatly improves the stability of heterologous proteins produced by A. niger.

Item Type:Article
Uncontrolled Keywords:Aspergillus niger, Cutinase
Subjects:T Technology > TP Chemical technology
Divisions:Chemical and Energy Engineering
ID Code:84458
Deposited By: Widya Wahid
Deposited On:11 Jan 2020 15:08
Last Modified:11 Jan 2020 15:08

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