The corpora cardiaca (CC) of the pneumorid grasshopper species Bullacris discolor contain at least one substance that causes hyperlipaemia in the migratory locust. Isolation of neuropeptides belonging to the adipokinetic hormone (AKH) family was achieved by single-step reversed-phase high performance liquid chromatography (RP-HPLC) of CC extracts and monitoring tryptophan fluorescence. The material of the bladder grasshopper showed three distinct fluorescence peaks with adipokinetic activity in the migratory locust. The peptide sequences were identified by Edman degradation after the N-terminal pyroglutamate residue had been cleaved off enzymatically, and the exact peptide masses were determined by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. Moreover, the assigned peptides were synthesised and natural and synthetic peptides were compared in their behaviour in RP-HPLC. B. discolor stores three AKH peptides in its CC: two of those are octapeptides, Schgr-AKH-II (pELNFSTGWamide) and Peram-CAH-II (pELTFTPNWamide), whereas the third peptide is a decapeptide, Phyle-CC (pELTFTPNWGSamide. The concentration of carbohydrates in the haemolymph of B. discolor is about 3 times higher than the lipid concentration. Upon injection with synthetic Schgr-AKH-II no adipokinetic or hypertrehalosaemic effect was measurable. A literature survey appears to indicate that an active role of these AKH peptides in substrate mobilisation is only overtly displayed in those caeliferan species that undertake well-defined flight periods.
Emp-AKH is a member of the large adipokinetic hormone (AKH) family of peptides. This peptide family appears to occur in the corpora cardiaca of all insect species and its members are involved in regulating substrate mobilisation. The secondary structure of Emp-AKH has been studied in the presence of sodium dodecyl sulfate micelles by comparing data obtained from Nuclear Magnetic Resonance and molecular dynamics simulations. The lowest energy conformer obtained in this study has a turn consisting of residues 5-8 and a tail consisting of the first five residues., Igor Z. Zubrzycki, Gerd Gäde, and Lit
We measured energy substrates in haemolymph and flight muscles of the large blister beetle Mylabris oculata at rest and after tethered, lift-generating flight. Flight of 1 min duration at an ambient temperature of 38-42°C did not effect a change in the concentration of lipids in the haemolymph, whereas a small, significant decrease in the concentrations of carbohydrates and a 3-fold larger one in the levels of proline were noted, as well as a concomitant increase in alanine. In the flight muscles, glycogen and proline concentrations were diminished slightly but significantly upon flight, whereas alanine levels were increased. Two hours of rest after a flight of 1 min completely reversed the metabolic situation in haemolymph and flight muscles to pre-flight levels. We could isolate two neuropeptides from the corpora cardiaca of M. oculata, which by retention time and mass analyses are characterised as the decapeptide Del-CC (pGlu-Leu-Asn-Phe-Ser-Pro-Asn-Trp-Gly-Asn-NH2) and the octapeptide Tem-HrTH (pGlu-Leu-Asn-PheSer-Pro-Asn-Trp-NH2) previously fully identified from the corpora cardiaca of the blister beetle, Decapotoma lunata (Gäde, 1995). Subsequently, it was unequivocally demonstrated that low doses of Del-CC and Tem-HrTH elicited increases in the concentration of proline and carbohydrates in the haemolymph of D. lunata and M. oculata, but did not change the concentration of lipids in both species. In conclusion, the two endogenous peptides are hypertrehalosaemic and hyperprolinaemic, thus very likely regulating the mobilisation of the two important flight substrates of blister beetles, namely carbohydrates and proline., Gerd Gäde, Lutz Auerswald, and Lit
Myotropic neuropeptides were isolated from the retrocerebral complex of the stick insect, Carausius morosus, by using three HPLC steps. Bioactivity during purification was measured by heterologous bioassays monitoring the contractions of the hyperneural muscle and hindgut of the American cockroach. Additionally, fractions not active in these bioassays were tested in a homologous bioassay evoking contractions of the hindgut of C. morosus. Peptide sequence analysis and mass spectrometry yielded the following structures: Pro-Phe-Cys-Asn-Ala-Phe-Thr-Gly-Cys-NH2 (CCAP), pGlu-Thr-Phe-Gln-Tyr-Ser-His-Gly-Trp-Thr-Asn-NH2 (His7-corazonin) and Asp-Glu-Gly-Gly-Thr-Gln-Tyr-Thr-Pro-Arg-Leu-NH2 (Cam-PK-1). These neuropeptides are the first myotropins isolated from C. morosus. The most bioactive compound in the homologous bioassay, the C. morosus-hindgut assay, was CCAP., Reinhard Predel, Roland Kellner, Gerd Gäde, and Lit
The presence of adipokinetic activity in crude extracts of corpora cardiaca (CC) from the butterfly (Vanessa cardui L., Nymphalidae) was demonstrated by bioassay and Mas-AKH was revealed as the major adipokinetic hormone (AKH) by use of two different technologies of sequence elucidation: HPLC separation of the peptide followed by Edman degradation and Q-TOF mass spectrometry. In contrast to the time- and material-consuming conventional methods of peptide purification and sequencing, substantial structural data of the peptide were confirmed - post factum - from one pCC (pair of CC) by Q-TOF mass spectrometry. Only males of our laboratory colony showed a significant lipid increase in the haemolymph after injection of either crude CC extract (1 pCC equivalent) or 10 pmol of synthetic peptide., Gabriele Köllisch, Peter D. Verhaert, Matthias W. Lorenz, Roland Kellner, Gerd Gäde, Klaus H. Hoffmann, and Lit
The corpora cardiaca (CC) of the two grasshopper species Zonocerus elegans (Pyrgomorphidae) and Lamarckiana sparrmani (Pamphagidae) contain (a) substance(s) that cause(s) hyperlipaemia in the migratory locust and hypertrehalosaemia in the American cockroach. Isolation of neuropeptides belonging to the adipokinetic hormone (AKH) family was achieved by single-step reversed-phase high performance liquid chromatography of CC extracts from both species and monitoring tryptophan fluorescence. The material of both species showed three distinct fluorescence peaks with adipokinetic activity in the migratory locust. The peptides were identifid by at least two of the following methods: (1) sequencing by Edman degradation, (2) sequencing by tandem fast atom bombardment mass spectrometry, (3) mass determination by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry and (4) co-elution of the native and synthetic peptides. Both species were found to have three AKH peptides stored in the CC, but unlike in other grasshoppers, none of those peptides were decapeptides. In Z. elegans the following three octapeptides occur: Schgr-AKH-II (pELNFSTGWamide), Peram-CAH-II (pELTFTPNWamide) and Phymo-AKH-III (pEINFTPWWamide), whereas L. sparrmani contains the octapeptides Grybi-AKH (pEVNFSTGWamide), Pyrap-AKH (pELNFTPNWamide) and also Phymo-AKH-III. Conspecific bioassays show no adipokinetic and only a weak (not significant) hypertrehalosaemic effect (in the pamphagid grasshopper). Some explanations are offered on the possible role of these peptides in the species investigated by interpreting their life style.