Effect of Pichia pastoris host strain on the properties of recombinant Aspergillus niger endoglucanase, EglB

Shazilah Kamaruddin; Nor Muhammad Mahadi; Rosli Md Illias; Osman Hassan; Suhaila Sulaiman; William Broughton; Izwan Bharudin; Farah Diba Abu Bakar; Abdul Munir Abdul Murad

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Effect of Pichia pastoris host strain on the properties of recombinant Aspergillus niger endoglucanase, EglB

Author: Shazilah Kamaruddin ^{1
,

2
,

3} ; Nor Muhammad Mahadi ⁴ ; Rosli Md Illias ⁵ ; Osman Hassan ^{6
,

7
,

8} ; Suhaila Sulaiman ⁹ ; William Broughton ¹⁰ ; Izwan Bharudin ^{1
,

2
,

3} ; Farah Diba Abu Bakar ^{1
,

2
,

3} ; Abdul Munir Abdul Murad ^{1
,

2
,

3}
Author Information

1. School of Biosciences &
2. Biotechnology, Faculty of Science &
3. Technology, Universiti Kebangsaan Malaysia, Bangi 43600 Selangor, Malaysia
4. Institute of Systems Biology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor, Malaysia
5. Department of Bioprocess Engineering, Faculty of Chemical and Natural Resources Engineering, Universiti Teknologi Malaysia, 81310 Skudai, Johor, Malaysia.
6. School of Chemistry &
7. Food Technology, Faculty of Science &
8. Technology, Universiti Kebangsaan Malaysia, Bangi, 43600 Selangor, Malaysia
9. Felda Global Ventures Research and Development Sdn Bhd, FGV Innovation Centre (Biotechnology), Bandar Enstek, 71760 Negeri Sembilan, Malaysia
10. BAM Federal Institute for Materials Research and Testing, Unter den Eichen 87, 12205 Berlin, Germany
Publication Type:Journal Article
Keywords: cellulase; endoglucanase; glycosylation; methanol utilisation phenotype; recombinant enzyme
From:Malaysian Journal of Microbiology 2018;14(6):554-562
CountryMalaysia
Language:English
Abstract: Aims:The methylotrophic yeast Pichia pastoris is widely used to express foreign proteins fused to secretion signals. As the effect of the expression host on the final protein product is unclear, we compared the properties of an endoglucanase (eglB of Aspergillus niger) expressed in two different P. pastoris strains.
Methodology and results:Full-length cDNA encoding endoglucanase of A. niger strain ATCC10574 was isolated and expressed in P. pastoris X33 (the methanol utilisation plus phenotype, Mut+) and P. pastoris GS115 (slow methanol utilisation, MutS). EglB-GS115 showed the highest activity and stability at 60 °C while EglB-X33 was most active at 50 °C. EglB-X33 was active towards other substrates such as arabinogalactan, guar gum and locust bean gum besides its specific substrate, carboxymethyl cellulose (CMC). However, EglB-GS115 was only active on CMC. The affinity of EglB-X33 towards CMC (Km = 7.5 mg/mL and specific activity 658 U/mg) was higher than that of EglB-GS115 (Km = 11.57 mg/mL, specific activity 144 U/mg).
Conclusion, significance and impact of study:Although eglB was cloned in the same expression vector (pPICZαC), two different characteristics of enzymes were recovered from the supernatant of the different hosts. Thus, expression of recombinant enzyme in different P. pastoris strains greatly affects the physical structure and biochemical properties of the enzyme.
Full text:20.2018my0488.pdf