Expression, purification and characterization of the Cry2Aa14 toxin from Bacillus thuringiensis subsp. kenyae
An indigenous strain HD-550 of Bacillus thuringiensis subsp. kenyae was found to be toxic to lepidopteran as well as dipteran insects. The cry2Aa gene (classified as cry2Aa14) from this isolate was cloned and expressed in Escherichia coli. Only a little amount of the expressed Cry2Aa14 protein was o...
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| Veröffentlicht in: | Toxicon (Oxford) 2009-09, Vol.54 (4), p.519-524 |
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| creator | Hire, Ramesh S. Makde, Ravindra D. Dongre, Tanaji K. D'souza, Stanislaus F. |
| description | An indigenous strain HD-550 of
Bacillus thuringiensis subsp.
kenyae was found to be toxic to lepidopteran as well as dipteran insects. The
cry2Aa gene (classified as
cry2Aa14) from this isolate was cloned and expressed in
Escherichia coli. Only a little amount of the expressed Cry2Aa14 protein was observed in soluble fraction under normal induction condition. The inclusions were non-toxic to test insects, whereas solubilized Cry2Aa14 was highly toxic to lepidopteran and dipteran insects. Cry2Aa14 protein was expressed as thioredoxin (trx) fusion protein for improving the yield of active protein. An enhancement of nearly 15% was observed in the yield of active Cry2Aa14. The TrxA–Cry2Aa14 protein purified from the solubilized fraction also showed toxicity profile similar to the wild-type protein. The LC
50 values of Cry2Aa14 and TrxA–Cry2Aa14 protein against
Spodoptera litura was 694 and 696
ng/cm
2, respectively, while for
Culex quinquefasciatus the LC
50 values were 894 and 902
ng/ml, respectively. The broad spectrum toxicity of the Cry2Aa14 thus indicates that this protein could be an important component in integrated pest management. Further, the trx tag clearly led to higher yield, which facilitates protein purification for biophysical and biochemical characterization. |
| doi_str_mv | 10.1016/j.toxicon.2009.05.022 |
| format | Article |
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Bacillus thuringiensis subsp.
kenyae was found to be toxic to lepidopteran as well as dipteran insects. The
cry2Aa gene (classified as
cry2Aa14) from this isolate was cloned and expressed in
Escherichia coli. Only a little amount of the expressed Cry2Aa14 protein was observed in soluble fraction under normal induction condition. The inclusions were non-toxic to test insects, whereas solubilized Cry2Aa14 was highly toxic to lepidopteran and dipteran insects. Cry2Aa14 protein was expressed as thioredoxin (trx) fusion protein for improving the yield of active protein. An enhancement of nearly 15% was observed in the yield of active Cry2Aa14. The TrxA–Cry2Aa14 protein purified from the solubilized fraction also showed toxicity profile similar to the wild-type protein. The LC
50 values of Cry2Aa14 and TrxA–Cry2Aa14 protein against
Spodoptera litura was 694 and 696
ng/cm
2, respectively, while for
Culex quinquefasciatus the LC
50 values were 894 and 902
ng/ml, respectively. The broad spectrum toxicity of the Cry2Aa14 thus indicates that this protein could be an important component in integrated pest management. Further, the trx tag clearly led to higher yield, which facilitates protein purification for biophysical and biochemical characterization.</description><identifier>ISSN: 0041-0101</identifier><identifier>EISSN: 1879-3150</identifier><identifier>DOI: 10.1016/j.toxicon.2009.05.022</identifier><identifier>PMID: 19486907</identifier><identifier>CODEN: TOXIA6</identifier><language>eng</language><publisher>Kidlington: Elsevier Ltd</publisher><subject>Animal poisons toxicology. Antivenoms ; Animals ; Bacillus thuringiensis ; Bacillus thuringiensis - genetics ; Bacillus thuringiensis - metabolism ; Bacillus thuringiensis subsp. kenyae ; Bacterial Proteins - chemistry ; Bacterial Proteins - genetics ; Bacterial Proteins - isolation & purification ; Bacterial Proteins - metabolism ; Biological and medical sciences ; Cloning, Molecular ; Cry2Aa protein ; Culex ; Culex quinquefasciatus ; Endotoxins - chemistry ; Endotoxins - genetics ; Endotoxins - isolation & purification ; Endotoxins - metabolism ; Escherichia coli ; Escherichia coli - genetics ; Helicoverpa armigera ; Hemolysin Proteins - chemistry ; Hemolysin Proteins - genetics ; Hemolysin Proteins - isolation & purification ; Hemolysin Proteins - metabolism ; Insecticides ; Lepidoptera ; Medical sciences ; Moths ; Recombinant Fusion Proteins - metabolism ; Solubility ; Spodoptera ; Spodoptera litura ; Thioredoxin fusion ; Toxicology</subject><ispartof>Toxicon (Oxford), 2009-09, Vol.54 (4), p.519-524</ispartof><rights>2009 Elsevier Ltd</rights><rights>2009 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c455t-1e1d6486e8fba7800aaea761d04717f1c0b978be5be81f341cfe55a150570a1e3</citedby><cites>FETCH-LOGICAL-c455t-1e1d6486e8fba7800aaea761d04717f1c0b978be5be81f341cfe55a150570a1e3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.toxicon.2009.05.022$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3550,27924,27925,45995</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=21798508$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/19486907$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Hire, Ramesh S.</creatorcontrib><creatorcontrib>Makde, Ravindra D.</creatorcontrib><creatorcontrib>Dongre, Tanaji K.</creatorcontrib><creatorcontrib>D'souza, Stanislaus F.</creatorcontrib><title>Expression, purification and characterization of the Cry2Aa14 toxin from Bacillus thuringiensis subsp. kenyae</title><title>Toxicon (Oxford)</title><addtitle>Toxicon</addtitle><description>An indigenous strain HD-550 of
Bacillus thuringiensis subsp.
kenyae was found to be toxic to lepidopteran as well as dipteran insects. The
cry2Aa gene (classified as
cry2Aa14) from this isolate was cloned and expressed in
Escherichia coli. Only a little amount of the expressed Cry2Aa14 protein was observed in soluble fraction under normal induction condition. The inclusions were non-toxic to test insects, whereas solubilized Cry2Aa14 was highly toxic to lepidopteran and dipteran insects. Cry2Aa14 protein was expressed as thioredoxin (trx) fusion protein for improving the yield of active protein. An enhancement of nearly 15% was observed in the yield of active Cry2Aa14. The TrxA–Cry2Aa14 protein purified from the solubilized fraction also showed toxicity profile similar to the wild-type protein. The LC
50 values of Cry2Aa14 and TrxA–Cry2Aa14 protein against
Spodoptera litura was 694 and 696
ng/cm
2, respectively, while for
Culex quinquefasciatus the LC
50 values were 894 and 902
ng/ml, respectively. The broad spectrum toxicity of the Cry2Aa14 thus indicates that this protein could be an important component in integrated pest management. Further, the trx tag clearly led to higher yield, which facilitates protein purification for biophysical and biochemical characterization.</description><subject>Animal poisons toxicology. Antivenoms</subject><subject>Animals</subject><subject>Bacillus thuringiensis</subject><subject>Bacillus thuringiensis - genetics</subject><subject>Bacillus thuringiensis - metabolism</subject><subject>Bacillus thuringiensis subsp. kenyae</subject><subject>Bacterial Proteins - chemistry</subject><subject>Bacterial Proteins - genetics</subject><subject>Bacterial Proteins - isolation & purification</subject><subject>Bacterial Proteins - metabolism</subject><subject>Biological and medical sciences</subject><subject>Cloning, Molecular</subject><subject>Cry2Aa protein</subject><subject>Culex</subject><subject>Culex quinquefasciatus</subject><subject>Endotoxins - chemistry</subject><subject>Endotoxins - genetics</subject><subject>Endotoxins - isolation & purification</subject><subject>Endotoxins - metabolism</subject><subject>Escherichia coli</subject><subject>Escherichia coli - genetics</subject><subject>Helicoverpa armigera</subject><subject>Hemolysin Proteins - chemistry</subject><subject>Hemolysin Proteins - genetics</subject><subject>Hemolysin Proteins - isolation & purification</subject><subject>Hemolysin Proteins - metabolism</subject><subject>Insecticides</subject><subject>Lepidoptera</subject><subject>Medical sciences</subject><subject>Moths</subject><subject>Recombinant Fusion Proteins - metabolism</subject><subject>Solubility</subject><subject>Spodoptera</subject><subject>Spodoptera litura</subject><subject>Thioredoxin fusion</subject><subject>Toxicology</subject><issn>0041-0101</issn><issn>1879-3150</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU1v1DAQhi0EotvCTwD5AqcmzCRx7JxQWZUPqRIXOFuOM6Ze8oWdoC6_Hq82gmNPlq1nxu_Mw9grhBwB63eHfJkevJ3GvABochA5FMUTtkMlm6xEAU_ZDqDCDBJ-wS5jPABAqZr6ObvAplJ1A3LHhtuHOVCMfhqv-bwG77w1S7pxM3bc3ptg7ELB_zk_To4v98T34VjcGKz4KcPIXZgG_sFY3_drTEBqM_7wNEYfeVzbOOf8J41HQy_YM2f6SC-384p9_3j7bf85u_v66cv-5i6zlRBLhoRdnRKScq2RCsAYMrLGDiqJ0qGFtpGqJdGSQldWaB0JYdLQQoJBKq_Y23PfOUy_VoqLHny01PdmpGmNuqxUhQXUj4IFyLoUKBMozqANU4yBnJ6DH0w4agR9EqIPehOiT0I0CJ2EpLrX2wdrO1D3v2ozkIA3G2CiNb0LZrQ-_uMKlI0SoBL3_sxR2ttvT0FHm1ZsqfOB7KK7yT8S5S8FG61q</recordid><startdate>20090915</startdate><enddate>20090915</enddate><creator>Hire, Ramesh S.</creator><creator>Makde, Ravindra D.</creator><creator>Dongre, Tanaji K.</creator><creator>D'souza, Stanislaus F.</creator><general>Elsevier Ltd</general><general>Elsevier</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7SS</scope><scope>7ST</scope><scope>7U6</scope><scope>7U7</scope><scope>C1K</scope><scope>8FD</scope><scope>FR3</scope><scope>KR7</scope></search><sort><creationdate>20090915</creationdate><title>Expression, purification and characterization of the Cry2Aa14 toxin from Bacillus thuringiensis subsp. kenyae</title><author>Hire, Ramesh S. ; Makde, Ravindra D. ; Dongre, Tanaji K. ; D'souza, Stanislaus F.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c455t-1e1d6486e8fba7800aaea761d04717f1c0b978be5be81f341cfe55a150570a1e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>Animal poisons toxicology. Antivenoms</topic><topic>Animals</topic><topic>Bacillus thuringiensis</topic><topic>Bacillus thuringiensis - genetics</topic><topic>Bacillus thuringiensis - metabolism</topic><topic>Bacillus thuringiensis subsp. kenyae</topic><topic>Bacterial Proteins - chemistry</topic><topic>Bacterial Proteins - genetics</topic><topic>Bacterial Proteins - isolation & purification</topic><topic>Bacterial Proteins - metabolism</topic><topic>Biological and medical sciences</topic><topic>Cloning, Molecular</topic><topic>Cry2Aa protein</topic><topic>Culex</topic><topic>Culex quinquefasciatus</topic><topic>Endotoxins - chemistry</topic><topic>Endotoxins - genetics</topic><topic>Endotoxins - isolation & purification</topic><topic>Endotoxins - metabolism</topic><topic>Escherichia coli</topic><topic>Escherichia coli - genetics</topic><topic>Helicoverpa armigera</topic><topic>Hemolysin Proteins - chemistry</topic><topic>Hemolysin Proteins - genetics</topic><topic>Hemolysin Proteins - isolation & purification</topic><topic>Hemolysin Proteins - metabolism</topic><topic>Insecticides</topic><topic>Lepidoptera</topic><topic>Medical sciences</topic><topic>Moths</topic><topic>Recombinant Fusion Proteins - metabolism</topic><topic>Solubility</topic><topic>Spodoptera</topic><topic>Spodoptera litura</topic><topic>Thioredoxin fusion</topic><topic>Toxicology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Hire, Ramesh S.</creatorcontrib><creatorcontrib>Makde, Ravindra D.</creatorcontrib><creatorcontrib>Dongre, Tanaji K.</creatorcontrib><creatorcontrib>D'souza, Stanislaus F.</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Environment Abstracts</collection><collection>Sustainability Science Abstracts</collection><collection>Toxicology Abstracts</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Civil Engineering Abstracts</collection><jtitle>Toxicon (Oxford)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Hire, Ramesh S.</au><au>Makde, Ravindra D.</au><au>Dongre, Tanaji K.</au><au>D'souza, Stanislaus F.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Expression, purification and characterization of the Cry2Aa14 toxin from Bacillus thuringiensis subsp. kenyae</atitle><jtitle>Toxicon (Oxford)</jtitle><addtitle>Toxicon</addtitle><date>2009-09-15</date><risdate>2009</risdate><volume>54</volume><issue>4</issue><spage>519</spage><epage>524</epage><pages>519-524</pages><issn>0041-0101</issn><eissn>1879-3150</eissn><coden>TOXIA6</coden><abstract>An indigenous strain HD-550 of
Bacillus thuringiensis subsp.
kenyae was found to be toxic to lepidopteran as well as dipteran insects. The
cry2Aa gene (classified as
cry2Aa14) from this isolate was cloned and expressed in
Escherichia coli. Only a little amount of the expressed Cry2Aa14 protein was observed in soluble fraction under normal induction condition. The inclusions were non-toxic to test insects, whereas solubilized Cry2Aa14 was highly toxic to lepidopteran and dipteran insects. Cry2Aa14 protein was expressed as thioredoxin (trx) fusion protein for improving the yield of active protein. An enhancement of nearly 15% was observed in the yield of active Cry2Aa14. The TrxA–Cry2Aa14 protein purified from the solubilized fraction also showed toxicity profile similar to the wild-type protein. The LC
50 values of Cry2Aa14 and TrxA–Cry2Aa14 protein against
Spodoptera litura was 694 and 696
ng/cm
2, respectively, while for
Culex quinquefasciatus the LC
50 values were 894 and 902
ng/ml, respectively. The broad spectrum toxicity of the Cry2Aa14 thus indicates that this protein could be an important component in integrated pest management. Further, the trx tag clearly led to higher yield, which facilitates protein purification for biophysical and biochemical characterization.</abstract><cop>Kidlington</cop><pub>Elsevier Ltd</pub><pmid>19486907</pmid><doi>10.1016/j.toxicon.2009.05.022</doi><tpages>6</tpages></addata></record> |
| fulltext | fulltext |
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| ispartof | Toxicon (Oxford), 2009-09, Vol.54 (4), p.519-524 |
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| source | MEDLINE; ScienceDirect Journals (5 years ago - present) |
| subjects | Animal poisons toxicology. Antivenoms Animals Bacillus thuringiensis Bacillus thuringiensis - genetics Bacillus thuringiensis - metabolism Bacillus thuringiensis subsp. kenyae Bacterial Proteins - chemistry Bacterial Proteins - genetics Bacterial Proteins - isolation & purification Bacterial Proteins - metabolism Biological and medical sciences Cloning, Molecular Cry2Aa protein Culex Culex quinquefasciatus Endotoxins - chemistry Endotoxins - genetics Endotoxins - isolation & purification Endotoxins - metabolism Escherichia coli Escherichia coli - genetics Helicoverpa armigera Hemolysin Proteins - chemistry Hemolysin Proteins - genetics Hemolysin Proteins - isolation & purification Hemolysin Proteins - metabolism Insecticides Lepidoptera Medical sciences Moths Recombinant Fusion Proteins - metabolism Solubility Spodoptera Spodoptera litura Thioredoxin fusion Toxicology |
| title | Expression, purification and characterization of the Cry2Aa14 toxin from Bacillus thuringiensis subsp. kenyae |
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