Molecular characterization of genes encoding trypsin-like enzymes from Aedes aegypti larvae and identification of digestive enzymes

• Published in: Gene 2011-12, Vol.489 (2), p.70-75
• Main Authors: Soares, Tatiane S., Watanabe, Renata M.O., Lemos, Francisco J.A., Tanaka, Aparecida S.
• Format: Article
• Language: English
• Subjects: adults; Aedes; Aedes - enzymology; Aedes - genetics; Aedes - metabolism; Aedes aegypti; Amino Acid Sequence; amino acids; Animals; BPTI; Digestion - physiology; Digestive System - enzymology; Digestive System - metabolism; Gene Expression; genes; instars; Larva - enzymology; Larva - genetics; Larva - metabolism; Larvae; midgut; Mosquito; Polymerase Chain Reaction; quantitative polymerase chain reaction; Sequence Alignment; Sequence Analysis, Protein; Serine protease inhibitor; Trypsin; Trypsin - genetics; Trypsin - metabolism
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/j.gene.2011.08.018

Trypsin-like enzymes play an important role in the Aedes aegypti digestive process. The trypsin-like enzymes present in adults were characterized previously, but little is known about trypsins in larvae. In the present work, we identified one of the trypsin enzymes from Ae. aegypti larval midgut using a library of trypsin gene fragments, which was the sequence known as AAEL005607 from the Ae. aegypti genome. Quantitative PCR analysis showed that AAEL005607 was transcribed in all larval instars, but it was not present in adult midgut. In order to confirm transcription data, the trypsin-like enzymes from 4th instar larvae of Ae. aegypti midgut were purified and sequenced. Purified trypsin showed identity with the amino-terminal sequence of AAEL005607, AAEL005609 and AAEL005614. These three trypsins have high amino acids identity, and could all be used as a template for the design of inhibitors. In conclusion, for the first time, digestive enzymes of 4th larval instar of Ae. aegypti were purified and characterized. The knowledge of digestive enzymes present in Ae. aegypti larvae may be helpful in the development of a larvicide.