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| Author(s): |
Adriana Rios Lopes
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | São Paulo. |
| Institution: | Universidade de São Paulo (USP). Conjunto das Químicas (IQ e FCF) (CQ/DBDCQ) |
| Defense date: | 1999-09-22 |
| Examining board members: |
Walter Ribeiro Terra;
Iolanda Midea Cuccovia;
Helena Bonciani Nader
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| Advisor: | Walter Ribeiro Terra |
| Abstract | |
Trypsins are serine endopeptidases that hydrolyze peptide bonds at the carboxyl side of positively charged residues: arginine and lysine. Mammalian trypsin preferentially cleaves the peptide bond formed by arginine. Site directed mutagenesis has shown that trypsin specificity is related to residues present at the primary specificity site and to structural determinants like two surface loops. Differences in trypsins specificity may be the cause of some insects be resistant to serine endopeptidases plant inhibitors and to Bacillus thuringiensis toxins. Trypsins are usual enzymes in insects and are very important to protein digestion. There are few studies dealing with insect trypsin specificity. They generally consist in analyses of fragments formed by the action of the enzyme on peptide chains like insulin β chain. As these studies were semi-quantitative, insect trypsin specificity requires a better characterization. This dissertation describes the purification of trypsins from Periplaneta americana, Tenebrio molitor, Musca domestica and Diatraea saccharalis and the characterization of the specificity of the subsites S1, S2, S3 e S1\' by the use of quenched fluorescence peptide substrates. The results showed that trypsins from the mentioned insects have different specificities, including the primary specificity. Thus, Diatraea saccharalis trypsin cleaves at Lys more efficiently than at Arg, whereas the eontrary is true for the other insects. The data also showed that trypsin subsites tend to beeome more hydrophobic as the insects are more evolved. (AU) | |