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We report for the first time the expression of multiple protease activities in the first instar larva (L1) of the flesh fly Oxysarcodexia thornax (Walker). Zymographic analysis of homogenates from freshly obtained L1 revealed a complex proteolytic profile ranging from 21.5 to 136 kDa. Although some activities were detected at pH 3.5 and 5.5, the optimum pH for most of the proteolytic activities was between pH 7.5 and 9.5. Seven of 10 proteases were completely inactivated by phenyl-methyl sulfonyl-fluoride, suggesting that main proteases expressed by L1 belong to serine proteases class. Complete inactivation of all enzymatic activities was obtained using N-p-Tosyl-L-phenylalanine chloromethyl ketone (100 µM), a specific inhibitor of chymotrypsin-like serine proteases.Keywords: Oxysarcodexia thornax -Sarcophagidae -chymotrypsin-like serine proteasesThe genus Oxysarcodexia belongs to the Sarcophagidae family (flesh flies) whose major biological feature is the ovo-larvipary (Pape 1996). Females may deposit first instar larvae directly onto the vertebrate host at predisposing sites (causing secondary myiasis), carcasses, decomposing organic matter, and dung (Panu et al. 2000). Genus Oxysarcodexia is widely distributed in the Neotropical regions (Lopes 1946), and in Brazil, O. thornax (Walker) is found in both urban and rural areas of several states. This species is abundant in the state of Rio de Janeiro, occurring preferentially in the summer season, with the population peak between January and February (Oliveira et al. 2002).In biological systems, proteases may carry out (i) a regulatory function by activation or inactivation of specific proteins via selective proteolysis, and (ii) a nonspecific proteolytic function involved in general protein hydrolysis. Such proteolytic mechanisms are highly regulated and are involved in a variety of physiologic processes, such as programmed cell death, stress responses (heat shock and anoxia), skeletal muscle atrophy, cellcell recognition, signal transduction and learning, morphogenesis, and photoreceptor light adaptation (Mykles 1998). Different classes of proteases such as cysteine proteases, metalloproteases and serino proteases have been described both in adults and larvae of several species of the Diptera (Han et al. 1997, Rosenfeld & Vanderberg 1998, Cho et al. 1999, Noriega et al. 2002, Vierstraete et al. 2003, Okuda et al. 2005, Fazito do Vale et al. 2007, Pires et al. 2007, Rodrigues et al. 2007). Despite the existence of information about morphology, taxonomy and ecology of this genus, as far as we know, no data has been published on the biochemical traits of O. thornax. In the present study, the proteolytic profile of O. thornax first instar larvae was investigated using zymographic analysis. O. thornax adults were collected as previously described (Oliveira et al. 2002) and females were kept in a 50 ml plastic vial for 24 h at room temperature. Following death of the females, abundant live first instar larvae (L1) were released and immediately collected. Larvae were ...

Two psychrophilic bacterial samples were isolated from King George Island soil, in Antarctica. The phylogenetic analysis based on the 16S rRNA (rrs) gene led to the correlation with the closest related isolates as Sporosarcina aquimarina (99%) and Algoriphagus antarcticus (99%), with query coverage of 99% and 98%, respectively. The spent culture media from both isolates displayed proteolytic activities detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis containing gelatin as protein substrate. Under the employed conditions, S. aquimarina showed a 55 kDa protease with the best activity detected at pH 7.0 and at 27°C. A. antarcticus also showed a single extracellular protease, however its molecular mass was around 90kDa and its best activity was detected at pH 9.0 and at 37°C. The proteases from both isolates were inhibited by 1,10-phenanthroline and EDTA, two metalloprotease inhibitors. This is the first record of protease detection in both species, and our results may contribute to broaden the basic knowledge of proteases from the Antarctica environment and may help prospecting future biotechnological applications of these enzymes.

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