Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1

Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1

The often-encountered problem such as protein degradation has driven various methods of cell lysis in obtaining recombinant protein post fermentation. In this paper, we compare methods such as homogenization, sonication, sonication with lysozyme and chemical lysis using B-PER reagent with lysozym...

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Main Authors: Mohd Arshad, Zatul Iffah, Amid, Azura, Othman, Muhd. Ezza Faiez
Format: Article
Language:English
Published: Universiti Teknologi Malaysia 2015
Subjects:
TP248.13 Biotechnology
Online Access:http://irep.iium.edu.my/46349/
http://irep.iium.edu.my/46349/
http://irep.iium.edu.my/46349/1/Cell_disruption.pdf
id iium-46349
recordtype eprints
spelling iium-463492016-05-16T00:57:50Z http://irep.iium.edu.my/46349/ Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1 Mohd Arshad, Zatul Iffah Amid, Azura Othman, Muhd. Ezza Faiez TP248.13 Biotechnology The often-encountered problem such as protein degradation has driven various methods of cell lysis in obtaining recombinant protein post fermentation. In this paper, we compare methods such as homogenization, sonication, sonication with lysozyme and chemical lysis using B-PER reagent with lysozyme to extract the recombinant bromelain from E. coli BL21-AI. The sonication process is found to be the most effective in releasing recombinant bromelain without any pre-treatment. To obtain the high quality of protein from sonication method, the influence of different extractant buffer was investigated including phosphate buffer saline (PBS), PBS containing cysteine and EDTA (PBS-CE), and sodium phosphate buffer containing cysteine and EDTA (EB-CE). The highest specific enzyme activity was obtained when it was extracted with EB-CE buffer. Under sodium dodecyl sulfate polyacrylamide gel electrophoresis, the recombinant bromelain showed protein band at 55kDa. In conclusion, the sonication method with extractant buffer containing 100mM phosphate buffer pH7.0 with 15 mM cysteine and 2 mM EDTA (EB-CE) was shown to give high specific activity of recombinant bromelain. Universiti Teknologi Malaysia 2015-12 Article PeerReviewed application/pdf en http://irep.iium.edu.my/46349/1/Cell_disruption.pdf Mohd Arshad, Zatul Iffah and Amid, Azura and Othman, Muhd. Ezza Faiez (2015) Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1. Jurnal Teknologi, 77 (24). pp. 83-87. ISSN 0127–9696 E-ISSN 2180–3722 http://www.jurnalteknologi.utm.my/index.php/jurnalteknologi/article/view/6712
repository_type Digital Repository
institution_category Local University
institution International Islamic University Malaysia
building IIUM Repository
collection Online Access
language English
topic TP248.13 Biotechnology
spellingShingle TP248.13 Biotechnology
Mohd Arshad, Zatul Iffah
Amid, Azura
Othman, Muhd. Ezza Faiez
Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1
description The often-encountered problem such as protein degradation has driven various methods of cell lysis in obtaining recombinant protein post fermentation. In this paper, we compare methods such as homogenization, sonication, sonication with lysozyme and chemical lysis using B-PER reagent with lysozyme to extract the recombinant bromelain from E. coli BL21-AI. The sonication process is found to be the most effective in releasing recombinant bromelain without any pre-treatment. To obtain the high quality of protein from sonication method, the influence of different extractant buffer was investigated including phosphate buffer saline (PBS), PBS containing cysteine and EDTA (PBS-CE), and sodium phosphate buffer containing cysteine and EDTA (EB-CE). The highest specific enzyme activity was obtained when it was extracted with EB-CE buffer. Under sodium dodecyl sulfate polyacrylamide gel electrophoresis, the recombinant bromelain showed protein band at 55kDa. In conclusion, the sonication method with extractant buffer containing 100mM phosphate buffer pH7.0 with 15 mM cysteine and 2 mM EDTA (EB-CE) was shown to give high specific activity of recombinant bromelain.
format Article
author Mohd Arshad, Zatul Iffah
Amid, Azura
Othman, Muhd. Ezza Faiez
author_facet Mohd Arshad, Zatul Iffah
Amid, Azura
Othman, Muhd. Ezza Faiez
author_sort Mohd Arshad, Zatul Iffah
title Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1
title_short Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1
title_full Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1
title_fullStr Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1
title_full_unstemmed Comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in E.coli BL21-A1
title_sort comparison of different cell disruption methods and cell extractant buffers for recombinant bromelain expressed in e.coli bl21-a1
publisher Universiti Teknologi Malaysia
publishDate 2015
url http://irep.iium.edu.my/46349/
http://irep.iium.edu.my/46349/
http://irep.iium.edu.my/46349/1/Cell_disruption.pdf
first_indexed 2023-09-18T21:05:59Z
last_indexed 2023-09-18T21:05:59Z
_version_ 1777410936358830080

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