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| Format: | Artículo científico |
| Language: | en |
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Universidad Nacional de Colombia
2010
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| Online Access: | https://www.redalyc.org/articulo.oa?id=77617808002 |
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Table of Contents:
- Preferential Protection of Domains II and III of Bacillus thuringiensis Cry1Aa Toxin by Brush Border Membrane Vesicles Syed-Rehan A. Hussain Álvaro M. Flórez Donald H. Dean Óscar Alzate Biología endotoxin Bacillus thuringiensis site directed mutagenesis The surface exposed Leucine 371 on loop 2 of domain II, in Cry1Aa toxin, was mutated to Lysine to generate the trypsin-sensitive mutant, L371K. Upon trypsin digestion L371K is cleaved into approximately 37 and 26 kDa fragments. These are separable on SDS-PAGE, but remain as a single molecule of 65 kDa upon purification by liquid chromatography. The larger fragment is domain I and a portion of domain II (amino acid residues 1 to 371). The smaller 26-kDa polypeptide is the remainder of domain II and domain III (amino acids 372 to 609). When the mutant toxin was treated with high dose of M. sexta gut juice both fragments were degraded. However, when incubated with M. sexta BBMV, the 26 kDa fragment (domains II and III) was preferentially protected from gut juice proteases. As previously reported, wild type Cry1Aa toxin was also protected against degradation by gut juice proteases when incubated with M. sexta BBMV. On the contrary, when mouse BBMV was added to the reaction mixture neither Cry1Aa nor L371K toxins showed resistance to M. sexta gut juice proteases and were degraded. Since the whole Cry1Aa toxin and most of the domain II and domain III of L371K are protected from proteases in the presence of BBMV of the target insect, we suggest that the insertion of the toxin into the membrane is complex and involves all three domains. 2010 artículo científico 0123-3475 https://www.redalyc.org/articulo.oa?id=77617808002 en http://www.redalyc.org/revista.oa?id=776 Revista Colombiana de Biotecnología application/pdf Universidad Nacional de Colombia Revista Colombiana de Biotecnología (Colombia) Num.2 Vol.XII