Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12

Psychrophilic basidiomycete yeast, Glaciozyma antarctica strain PI12, was shown to be a protease-producer. Isolation of the PI12 protease gene from genomic and mRNA sequences allowed determination of 19 exons and 18 introns. Full-length cDNA of PI12 protease gene was amplified by rapid amplification...

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Main Authors:	Alias, Norsyuhada, Ahmad Mazian, Mu’adz, Salleh, Abu Bakar, Basri, Mahiran, Raja Abd. Rahman, Raja Noor Zaliha
Format:	Article
Language:	English
Published:	Hindawi Publishing Corporation 2014
Subjects:	QR Microbiology
Online Access:	http://irep.iium.edu.my/53050/ http://irep.iium.edu.my/53050/ http://irep.iium.edu.my/53050/ http://irep.iium.edu.my/53050/1/197938.pdf

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recordtype	eprints
spelling	iium-530502016-12-01T05:35:30Z http://irep.iium.edu.my/53050/ Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12 Alias, Norsyuhada Ahmad Mazian, Mu’adz Salleh, Abu Bakar Basri, Mahiran Raja Abd. Rahman, Raja Noor Zaliha QR Microbiology Psychrophilic basidiomycete yeast, Glaciozyma antarctica strain PI12, was shown to be a protease-producer. Isolation of the PI12 protease gene from genomic and mRNA sequences allowed determination of 19 exons and 18 introns. Full-length cDNA of PI12 protease gene was amplified by rapid amplification of cDNA ends (RACE) strategy with an open reading frame (ORF) of 2892 bp, coded for 963 amino acids. PI12 protease showed low homology with the subtilisin-like protease from fungus Rhodosporidium toruloides (42% identity) and no homology to other psychrophilic proteases. The gene encoding mature PI12 protease was cloned into Pichia pastoris expression vector, pPIC9, and positioned under the induction of methanol-alcohol oxidase (AOX) promoter. The recombinant PI12 protease was efficiently secreted into the culture medium driven by the Saccharomyces cerevisiae α-factor signal sequence. The highest protease production (28.3 U/ml) was obtained from P. pastoris GS115 host (GpPro2) at 20°C after 72 hours of postinduction time with 0.5% (v/v) of methanol inducer. The expressed protein was detected by SDS-PAGE and activity staining with a molecular weight of 99 kDa. Hindawi Publishing Corporation 2014-06-30 Article PeerReviewed application/pdf en http://irep.iium.edu.my/53050/1/197938.pdf Alias, Norsyuhada and Ahmad Mazian, Mu’adz and Salleh, Abu Bakar and Basri, Mahiran and Raja Abd. Rahman, Raja Noor Zaliha (2014) Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12. Enzyme Research, 2014. pp. 1-20. ISSN 20900406 https://www.hindawi.com/journals/er/2014/197938/ http://dx.doi.org/10.1155/2014/197938
repository_type	Digital Repository
institution_category	Local University
institution	International Islamic University Malaysia
building	IIUM Repository
collection	Online Access
language	English
topic	QR Microbiology
spellingShingle	QR Microbiology Alias, Norsyuhada Ahmad Mazian, Mu’adz Salleh, Abu Bakar Basri, Mahiran Raja Abd. Rahman, Raja Noor Zaliha Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12
description	Psychrophilic basidiomycete yeast, Glaciozyma antarctica strain PI12, was shown to be a protease-producer. Isolation of the PI12 protease gene from genomic and mRNA sequences allowed determination of 19 exons and 18 introns. Full-length cDNA of PI12 protease gene was amplified by rapid amplification of cDNA ends (RACE) strategy with an open reading frame (ORF) of 2892 bp, coded for 963 amino acids. PI12 protease showed low homology with the subtilisin-like protease from fungus Rhodosporidium toruloides (42% identity) and no homology to other psychrophilic proteases. The gene encoding mature PI12 protease was cloned into Pichia pastoris expression vector, pPIC9, and positioned under the induction of methanol-alcohol oxidase (AOX) promoter. The recombinant PI12 protease was efficiently secreted into the culture medium driven by the Saccharomyces cerevisiae α-factor signal sequence. The highest protease production (28.3 U/ml) was obtained from P. pastoris GS115 host (GpPro2) at 20°C after 72 hours of postinduction time with 0.5% (v/v) of methanol inducer. The expressed protein was detected by SDS-PAGE and activity staining with a molecular weight of 99 kDa.
format	Article
author	Alias, Norsyuhada Ahmad Mazian, Mu’adz Salleh, Abu Bakar Basri, Mahiran Raja Abd. Rahman, Raja Noor Zaliha
author_facet	Alias, Norsyuhada Ahmad Mazian, Mu’adz Salleh, Abu Bakar Basri, Mahiran Raja Abd. Rahman, Raja Noor Zaliha
author_sort	Alias, Norsyuhada
title	Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12
title_short	Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12
title_full	Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12
title_fullStr	Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12
title_full_unstemmed	Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12
title_sort	molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica pi12
publisher	Hindawi Publishing Corporation
publishDate	2014
url	http://irep.iium.edu.my/53050/ http://irep.iium.edu.my/53050/ http://irep.iium.edu.my/53050/ http://irep.iium.edu.my/53050/1/197938.pdf
first_indexed	2023-09-18T21:15:07Z
last_indexed	2023-09-18T21:15:07Z
_version_	1777411510395469824

Molecular cloning and optimization for high level expression of cold-adapted serine protease from antarctic yeast glaciozyma antarctica PI12

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