Browsing by Author "Izwan Bharudin"
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Publication Effect of Pichia pastoris host strain on the properties of recombinant Aspergillus niger endoglucanase, EglB(Malaysian Society for Microbiology, 2018) ;Shazilah Kamaruddin ;Nor Muhammad Mahadi ;Rosli Md Illias ;Osman Hassan ;Suhaila Sulaiman ;William Broughton ;Izwan Bharudin ;Farah Diba Abu BakarAbdul Munir Abdul MuradAims: 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-1 and specific activity 658 U.mg-1) was higher than that of EglB-GS115 (Km = 11.57 mg.mL-1, specific activity 144 U.mg-1). 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. Keywords: cellulase, endoglucanase, glycosylation, methanol utilisation phenotype, recombinant enzyme - Some of the metrics are blocked by yourconsent settings
Publication The Glaciozyma Antarctica Genome Reveals An Array Of Systems That Provide Sustained Responses Towards Temperature Variations In A Persistently Cold Habitat(Public Library of Science (PLOS), 2018) ;Mohd Firdaus-Raih ;Noor Haza Fazlin Hashim ;Izwan Bharudin ;Mohd Faizal Abu Bakar ;Kie Kyon Huang ;Halimah Alias ;Bernard K. B. Lee ;Mohd Noor Mat Isa ;Shuhaila Mat-Sharani ;Suhaila Sulaiman ;Lih Jinq Tay ;Radziah Zolkefli ;Yusuf Muhammad Noor ;Douglas Sie Nguong Law ;Siti Hamidah Abdul Rahman ;Rosli Md-Illias ;Farah Diba Abu Bakar ;Nazalan Najimudin ;Abdul Munir Abdul MuradNor Muhammad MahadiExtremely low temperatures present various challenges to life that include ice formation and effects on metabolic capacity. Psyhcrophilic microorganisms typically have an array of mechanisms to enable survival in cold temperatures. In this study, we sequenced and analysed the genome of a psychrophilic yeast isolated in the Antarctic region, Glaciozyma antarctica. The genome annotation identified 7857 protein coding sequences. From the genome sequence analysis we were able to identify genes that encoded for proteins known to be associated with cold survival, in addition to annotating genes that are unique to G. antarctica. For genes that are known to be involved in cold adaptation such as anti-freeze proteins (AFPs), our gene expression analysis revealed that they were differentially transcribed over time and in response to different temperatures. This indicated the presence of an array of adaptation systems that can respond to a changing but persistent cold environment. We were also able to validate the activity of all the AFPs annotated where the recombinant AFPs demonstrated anti-freeze capacity. This work is an important foundation for further collective exploration into psychrophilic microbiology where among other potential, the genes unique to this species may represent a pool of novel mechanisms for cold survival.