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

Nurhaida Kamaruddin, Reginald Storms, Nor Muhammad Mahadi, Rosli Md Illias, Farah Diba Abu Bakar, Abdul Munir Abd. Murad

Research output: Contribution to journalArticle

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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.

Original languageEnglish
Pages (from-to)3327-3338
Number of pages12
JournalArabian Journal for Science and Engineering
Volume43
Issue number7
DOIs
Publication statusPublished - 1 Jul 2018

Fingerprint

Aspergillus niger
Protein Stability
Peptide Hydrolases
Genes
Phyllachorales
Glucan 1,4-alpha-Glucosidase
Proteins
Transcription Factors
cutinase
Genome
Polymerase Chain Reaction

Keywords

  • Aspergillus niger
  • Cutinase
  • Extracellular proteases
  • Transcription factor

ASJC Scopus subject areas

  • General

Cite this

Reduction of Extracellular Proteases Increased Activity and Stability of Heterologous Protein in Aspergillus niger. / Kamaruddin, Nurhaida; Storms, Reginald; Mahadi, Nor Muhammad; Illias, Rosli Md; Abu Bakar, Farah Diba; Abd. Murad, Abdul Munir.

In: Arabian Journal for Science and Engineering, Vol. 43, No. 7, 01.07.2018, p. 3327-3338.

Research output: Contribution to journalArticle

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