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 lysozyme t...

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Main Authors: Zatul Iffah, Mohd Arshad, Azura, Amid, Muhd. Ezza Faiez, Othman
Format: Article
Language:English
English
Published: Penerbit Universiti Teknologi Malaysia 2015
Subjects:
TP Chemical technology
Online Access:http://umpir.ump.edu.my/id/eprint/11859/
http://umpir.ump.edu.my/id/eprint/11859/
http://umpir.ump.edu.my/id/eprint/11859/1/Comparison%20Of%20Different%20Cell%20Disruption%20Methods%20And%20Cell%20Extractant%20Buffers%20For%20Recombinant%20Bromelain%20Expressed%20in%20E.COLI%20BL21-A1.pdf
http://umpir.ump.edu.my/id/eprint/11859/7/Comparison%20of%20Different%20Cell%20Disruption%20Methods%20and%20Cell%20Extractant%20Buffers%20for%20Recombinant%20Bromelain%20Expressed%20in%20E-coli%20BL21-A1.pdf
id ump-11859
recordtype eprints
spelling ump-118592016-04-21T07:50:32Z http://umpir.ump.edu.my/id/eprint/11859/ Comparison of Different Cell Disruption Methods And Cell Extractant Buffers For Recombinant Bromelain Expressed in E.COLI BL21-A1 Zatul Iffah, Mohd Arshad Azura, Amid Muhd. Ezza Faiez, Othman TP Chemical technology 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. Penerbit Universiti Teknologi Malaysia 2015 Article PeerReviewed application/pdf en http://umpir.ump.edu.my/id/eprint/11859/1/Comparison%20Of%20Different%20Cell%20Disruption%20Methods%20And%20Cell%20Extractant%20Buffers%20For%20Recombinant%20Bromelain%20Expressed%20in%20E.COLI%20BL21-A1.pdf application/pdf en http://umpir.ump.edu.my/id/eprint/11859/7/Comparison%20of%20Different%20Cell%20Disruption%20Methods%20and%20Cell%20Extractant%20Buffers%20for%20Recombinant%20Bromelain%20Expressed%20in%20E-coli%20BL21-A1.pdf Zatul Iffah, Mohd Arshad and Azura, Amid and Muhd. Ezza Faiez, Othman (2015) Comparison of Different Cell Disruption Methods And Cell Extractant Buffers For Recombinant Bromelain Expressed in E.COLI BL21-A1. Jurnal Teknologi (Sciences and Engineering), 77 (24). pp. 83-87. ISSN 0127-9696 (print); 2180-3722 (online) http://www.jurnalteknologi.utm.my/index.php/jurnalteknologi/article/view/6712
repository_type Digital Repository
institution_category Local University
institution Universiti Malaysia Pahang
building UMP Institutional Repository
collection Online Access
language English
English
topic TP Chemical technology
spellingShingle TP Chemical technology
Zatul Iffah, Mohd Arshad
Azura, Amid
Muhd. Ezza Faiez, Othman
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 Zatul Iffah, Mohd Arshad
Azura, Amid
Muhd. Ezza Faiez, Othman
author_facet Zatul Iffah, Mohd Arshad
Azura, Amid
Muhd. Ezza Faiez, Othman
author_sort Zatul Iffah, Mohd Arshad
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 Penerbit Universiti Teknologi Malaysia
publishDate 2015
url http://umpir.ump.edu.my/id/eprint/11859/
http://umpir.ump.edu.my/id/eprint/11859/
http://umpir.ump.edu.my/id/eprint/11859/1/Comparison%20Of%20Different%20Cell%20Disruption%20Methods%20And%20Cell%20Extractant%20Buffers%20For%20Recombinant%20Bromelain%20Expressed%20in%20E.COLI%20BL21-A1.pdf
http://umpir.ump.edu.my/id/eprint/11859/7/Comparison%20of%20Different%20Cell%20Disruption%20Methods%20and%20Cell%20Extractant%20Buffers%20for%20Recombinant%20Bromelain%20Expressed%20in%20E-coli%20BL21-A1.pdf
first_indexed 2023-09-18T22:12:52Z
last_indexed 2023-09-18T22:12:52Z
_version_ 1777415144306900992

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