INSECT CHYMOTRYPSINS: CHLOROMETHYL KETONE INACTIVATION AND SUBSTRATE SPECIFICITY RELATIVE TO POSSIBLE COEVOLUTIONAL ADAPTATION OF INSECTS AND PLANTS

Insect digestive chymotrypsins are present in a large variety of insect orders but their substrate specificity still remains unclear. Ewer insect chymotrypsins from 3 different insect orders (Dictyoptera, Coleoptera and two Lepidoptera) were isolated using affinity chromatography. Enzymes presented molecular masses in the range of 20 to 31 kDa and pH optima in the range of 7.5 to 10.0. Kinetic characterization. using different, colorimetric and fluorescent substrates indicated that insect chymotrypsins differ from, bovine chymotrypsin in their primary specificity toward small substrates (like N-benzoyl-L-Tyr p-nitroanilide) rather than on their preference for large substrates (exemplified by Succynil-Ala-Ala-Pro-Phe P-nitroanilide). Chloromethyl ketones (TPCK, N-alpha-tosyl-L-Phe chloromethyl ketone and Z-GGF-CK, N-carbobenzoxy-Gly-Gly-phe-CK) inactivated all chymotrypsins legated. Inactivation rates follow apparent first-order kinetics with variable second order rates (TPCK, 42 to 130 M(-1)s(-1); Z-GGF-CK, 150 to 450 M(-1)s(-1) that may be remarkably low for S. frugiperda chymotrypsin (TPCK, 6 M(-1)s(-1); Z-GGF-CK, 6.1 M(-1) s(-1)). Homology modelling and sequence alignment showed that. in lepidopteran chymotrypsins, differences in the amino acid residues in the neighborhood of the catalytic His 57 may affect its pKa, value. This is Proposed as the cause of the decrease in His 57 reactivity toward chloromethyl ketones. Such amino acid replacement in the active site is proposed. to be an adaptation to the presence of dietary ketones. (C) 2009 Wiley Periodicals, Inc.

Subsite substrate specificity of midgut insect chymotrypsins

Insect chymotrypsins are distinctively sensitive to plant protein inhibitors, suggesting that they differ in subsite architecture and hence in substrate specificities. Purified digestive chymotrypsins from insects of three different orders were assayed with internally quenched fluorescent oligopeptides with three different amino acids at P1 (Tyr, Phe, and Leu) and 13 amino acid replacements in positions P1`, P2, and P3. The binding energy (Delta G(s), calculated from Km values) and the activation energy (Delta G(T)(double dagger), determined from k(cat)/K-m values) were calculated. The hydrophobicities of each subsite were calculated from the efficiency of hydrolysis of the different amino acid replacements at that subsite. The results showed that except for S1, the other subsites (S2, S3, and S1`) vary among chymotrypsins. This result contrasts with insect trypsin data that revealed a trend along evolution, putatively associated with resistance to plant inhibitors. In spite of those differences, the data suggested that in lepidopteran chymotrypsins S2 and S1` bind the substrate ground state, whereas only S1` binds the transition state, supporting aspects of the present accepted mechanism of catalysis. 2008 Elsevier Ltd. All rights reserved.

Is water extraction a threat to insect populations in intermittently-flowing streams?

Stream insects use a variety of refuges to survive drought that vary in their vulnerability to water extraction. We surveyed drought refuges in 16 ephemeral streams with different flow regimes. Perennial sections were the most important drought refuge, but are the most likely to disappear when water is extracted.

Insect-inspired robotic homing

Many animals, including insects, successfully engage in visual homing. We describe a system that allows a mobile robot to home. Specifically, we propose a simple, yet robust, homing scheme that only relies upon the observation of the bearings of visible landmarks. However, this can easily be extended to include other visual cues. The homing algorithm allows a mobile robot to home incrementally by moving in such a way as to gradually reduce the discrepancy between the current view and the view obtained from the home position. Both simulation and mobile robot experiments are used to demonstrate the feasibility of the approach.

Insect based navigation and its applications to the autonomous control of mobile robots

Giving robots the ability to autonomously move around in various real-world environments has been a major goal of AI (artificial intelligence) for quite some time. To this end it is vital for robots to be able to perceive their surroundings in 3D; they must be able to estimate the range of obstacles in their path.

Animals navigate through various uncontrolled environments with seemingly little effort. Flying insects, especially, are quite adept at manoeuvring in complex, unpredictable and possibly hostile and hazardous environments.

In this paper it is shown that very simple motion cues, inspired by the visual navigation of flying insects, can be used to provide a mobile robot with the ability to successfully traverse a corridor environment. Equipping an autonomous mobile robot with the ability to successfully navigate real-word environments (in real-time) constitutes a major challenge for AI and robotics. It is in this area that insect based navigation has something to offer.

Insect inspired behaviours for the autonomous control of mobile robots

Animals navigate through various uncontrolled environments with seemingly little effort. Flying insects, especially, are quite adept at manoeuvring in complex, unpredictable and possibly hostile environments. Through both simulation and real-world experiments, we demonstrate the feasibility of equipping a mobile robot with the ability to navigate a corridor environment, in real time, using principles based on insect-based visual guidance. In particular we have used the bees’ navigational strategy of measuring object range in terms of image velocity. We have also shown the viability and usefulness of various other insect behaviours: (i) keeping walls equidistant, (ii) slowing down when approaching an object, (iii) regulating speed according to tunnel width, and (iv) using visual motion as a measure of distance travelled.

An insect-based approach to robotic homing

Many animals, including insects, successfully engage in visual homing. We describe a system that allows a mobile robot to home. Specifically we propose a simple extension to our original homing scheme which significantly improves its performance by incorporating a richer view of the environment. The addition of landmark apparent-size cues assists homing by providing a more robust homing vector as well as providing a simple and effective method of reinforcing landmark avoidance. The homing algorithm allows a mobile robot to incrementally home by moving in such a way as to gradually reduce the discrepancy between the current view and the view obtained from the home position. Both simulation and mobile robot experiments are used to demonstrate the feasibility of this approach. By matching the bearings of features extracted from panoramic views and using a vector summation technique to compute a homing vector we are able to provide a simple, parsimonious and robust robotic homing algorithm.

Subfossils of extinct and extant species of Simuliidae (Diptera) from Austral and Cook Islands (Polynesia): anthropogenic extirpation of an aquatic insect?

Subfossil head capsules of Simuliidae larvae have been recovered from swamps on Tubuai and Raivavae of the Austral Islands, and Atiu and Mangaia of the southern Cook Islands. For Tubuai and Raivavae it is likely that the simuliids are extinct, but a single simuliid species is extant on nearby Rurutu. For Atiu and Mangaia, extant simuliids have not been reported, but are known on Rarotonga. Well-preserved head capsules indicate that the Cook Islands subfossils are those of Simulium (Inseliellum) teruamanga Craig and Craig, 1986. For the Austral Islands, the simuliid from Tubuai is considered a variant of Simulium (Inseliellum) rurutuense Craig and Joy, 2000. That from Raivavae is morphologically distinct and is described here as a new species, Simulium (Inseliellum) raivavaense Craig and Porch. Humans arrived in Eastern Polynesia ca. 1,000 years ago resulting in the widespread destruction of lowland forest and conversion of wetlands to agriculture with implied consequences for the indigenous biota of these habitats. Here we consider that one such result was loss of freshwater aquatic biodiversity.

Sex differences in insect immune function : a consequence of diet choice?

Males often have reduced immune function compared to females but the proximate mechanisms underlying this taxonomically widespread pattern are unclear. Because immune function is resource-dependent and sexes may have different nutritional requirements, we hypothesized that sexual dimorphism in immune function may arise from differential nutrient intake (acquisition hypothesis). To test this hypothesis, we examined patterns of phenoloxidase (PO) activity in relation to nutrient consumption in Queensland fruit flies (Q-flies). In the first experiment, flies were allowed to choose their preferred nutrient intake. Compared with males, female Q-flies had higher PO activity, consumed more calories, and preferred a higher protein:carbohydrate (P:C) diet, suggesting that differential acquisition could explain sex differences. In the second experiment, we restricted flies to one of 12 diets varying in protein and carbohydrate concentrations and mapped PO activity for each sex onto a nutritional landscape. Counter to our hypothesis, females had higher PO activity than males at any given level of nutrient intake. Both carbohydrate and protein intake affected PO activity in females but only protein affected PO activity in males. Our results indicate that sex differences in Q-fly immune function are not solely explained by sex differences in nutrient intake, although nutrition does contribute to the magnitude of these sex differences.