Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus
Commercially available bromelain is prepared by performing a tedious and costly purification method that provides multiple degrees of purified bromelain. In the current study, a gene encoding stem bromelain from Ananas comosus was amplified using polymerase chain reaction (PCR). This bromelain gene...
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ELSEVIER
2011
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| Online Access: | http://irep.iium.edu.my/2374/ http://irep.iium.edu.my/2374/1/Bromelain_cloning.pdf |
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| author | Amid, Azura Ismail, Nurul Azira Yusof, Faridah Mohd. Salleh, Hamzah |
| author_facet | Amid, Azura Ismail, Nurul Azira Yusof, Faridah Mohd. Salleh, Hamzah |
| author_sort | Amid, Azura |
| building | IIUM Repository |
| collection | Online Access |
| description | Commercially available bromelain is prepared by performing a tedious and costly purification method that provides multiple degrees of purified bromelain. In the current study, a gene encoding stem bromelain from Ananas comosus was amplified using polymerase chain reaction (PCR). This bromelain gene was initially cloned into pENTR/TEV/D-TOPO before being sub-cloned into the expression vector pDEST17. DNA sequencing of the amplified products exhibited a high level of homology to the corresponding gene from the NCBI public database. Protein expression was conducted in the Escherichia coli, BL21-AI. The recombinant bromelain was then purified in a single step using a Ni-NTA spin column, an example of immobilized metal affinity chromatography. Purified recombinant bromelain was detected by Western blotting. In addition, the purified enzyme exhibited hydrolytic activity towards gelatin and a synthetic substrate, LNPE. The purified recombinant bromelain exhibited optimum activity at pH 4.6 and 45oC. |
| first_indexed | 2025-11-14T14:23:05Z |
| format | Article |
| id | iium-2374 |
| institution | International Islamic University Malaysia |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-14T14:23:05Z |
| publishDate | 2011 |
| publisher | ELSEVIER |
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| spelling | iium-23742021-07-14T01:42:35Z http://irep.iium.edu.my/2374/ Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus Amid, Azura Ismail, Nurul Azira Yusof, Faridah Mohd. Salleh, Hamzah Q Science (General) Commercially available bromelain is prepared by performing a tedious and costly purification method that provides multiple degrees of purified bromelain. In the current study, a gene encoding stem bromelain from Ananas comosus was amplified using polymerase chain reaction (PCR). This bromelain gene was initially cloned into pENTR/TEV/D-TOPO before being sub-cloned into the expression vector pDEST17. DNA sequencing of the amplified products exhibited a high level of homology to the corresponding gene from the NCBI public database. Protein expression was conducted in the Escherichia coli, BL21-AI. The recombinant bromelain was then purified in a single step using a Ni-NTA spin column, an example of immobilized metal affinity chromatography. Purified recombinant bromelain was detected by Western blotting. In addition, the purified enzyme exhibited hydrolytic activity towards gelatin and a synthetic substrate, LNPE. The purified recombinant bromelain exhibited optimum activity at pH 4.6 and 45oC. ELSEVIER 2011-09 Article PeerReviewed application/pdf en http://irep.iium.edu.my/2374/1/Bromelain_cloning.pdf Amid, Azura and Ismail, Nurul Azira and Yusof, Faridah and Mohd. Salleh, Hamzah (2011) Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus. Process Biochemistry, 46 (12). pp. 2232-2239. ISSN 1359-5113 http://www.elsevier.com/wps/find/journaldescription.cws_home/422857/description#description |
| spellingShingle | Q Science (General) Amid, Azura Ismail, Nurul Azira Yusof, Faridah Mohd. Salleh, Hamzah Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| title | Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| title_full | Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| title_fullStr | Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| title_full_unstemmed | Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| title_short | Expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| title_sort | expression, purification, and characterization of a recombinant stem bromelain from ananas comosus |
| topic | Q Science (General) |
| url | http://irep.iium.edu.my/2374/ http://irep.iium.edu.my/2374/ http://irep.iium.edu.my/2374/1/Bromelain_cloning.pdf |