► CAP2b analogs were synthesized with (Z)-alkene, and (E)-alkene components. ► These components lock in a cis- or trans-Proline orientation, respectively. ► They were tested for antidiuretic activity ...on stink bug Malpighian tubules. ► Only the trans-Pro analog had antidiuretic activity. ► This is consistent with the Pro in CAP2b adopting a trans orientation.
The CAP2b neuropeptide family plays an important role in the regulation of the processes of diuresis and/or antidiuresis in a variety of insects. While Manse-CAP2b (pELYAFPRV-NH2) and native CAP2bs elicit diuretic activity in a number of species of flies, native CAP2b sequences have been shown to elicit antidiuretic activity in the kissing bug Rhodnius prolixus and the green stink bug Acrosternum hilare, the latter being an important pest of cotton and soybean in the southern United States. Analogs of CAP2b containing either a (Z)-alkene, cis-Pro or an (E)-alkene, trans-Pro isosteric component were synthesized and evaluated in an in vitro stink bug diuretic assay, which involved measurement of fluid secretion by Malpighian tubules isolated from A. hilare. The conformationally constrained trans-Pro analog demonstrated statistically significant antidiuretic activity, whereas the cis-Pro analog failed to elicit activity. The results are consistent with the adoption of a trans orientation for the Pro in CAP2b neuropeptides during interaction with receptors associated with the antidiuretic process in the stink bug. In addition, the results are further consistent with a theory of ligand-receptor coevolution between the CAP2b and pyrokinin/PBAN neuropeptide classes, both members of the ‘-PRXamide’ superfamily. This work further identifies a scaffold with which to design mimetic CAP2b analogs as potential leads in the development of environmentally favorable pest management agents capable of disrupting CAP2b-regulated diuretic/antidiuretic functions.
Insect neuropeptides of the insect kinin class share a common C-terminal pentapeptide sequence F1X1²X2³W4G5-NH2 (X2³ = P, S, A) and regulate such critical physiological processes as water balance and ...digestive enzyme release. Analogs of the insect kinin class, in which the critical residues of F1, P3, and W4 were replaced with β3-amino acid or their β2-homo-amino acid variants, have been synthesized by the solid phase peptide strategy. The resulting single- and double-replacement analogs were evaluated in an insect diuretic assay and enzyme digestion trials. Analogs modified in the core P3 position produce a potent and efficacious diuretic response that is not significantly different from that obtained with the endogenous achetakinin peptides. The analogs also demonstrate enhanced resistance to hydrolysis by ACE and NEP, endopeptidases that inactivate the natural insect neuropeptides. This paper describes the first instance of β-amino acids analogs of an arthropod peptide that demonstrate significant bioactivity and resistance to peptidase degradation.
The pyrokinin/pheromone biosynthesis activating neuropeptide (PK/PBAN) family plays a multifunctional role in an array of important physiological processes in a variety of insects. An active core ...analog containing an (
E)-alkene,
trans-Pro isosteric component was evaluated in four disparate PK/PBAN bioassays in four different insect species. These bioassays include pheromone biosynthesis in the moth
Heliothis peltigera, melanization in the larval
Spodoptera littoralis, pupariation acceleration in the larval fly
Neobellieria bullata, and hindgut contraction in the cockroach
Leucophaea maderae. The conformationally constrained analog demonstrated activity equivalent to parent PK/PBAN peptides of equal length in all four PK/PBAN bioassays, and matched and/or approached the activity of peptides of natural length in three of them. In the melanization bioassay, the constrained analog exceeded the efficacy (maximal response) of the natural PBAN1-33 by a factor of 2 (at 1
nmol). The results provide strong evidence for the orientation of Pro and the core conformation adopted by PK/PBAN neuropeptides during interaction with receptors associated with a range of disparate PK/PBAN bioassays. The work further identifies a scaffold with which to design mimetic PK/PBAN analogs as potential leads in the development of environmentally favorable pest management agents capable of disrupting PK/PBAN-regulated systems.
The pyrokinin/pheromone biosynthesis activating neuropeptide (PK/PBAN) family plays a multifunctional role in an array of important physiological processes in insects, including regulation of sex ...pheromone biosynthesis in moths. A cyclic PK/PBAN analog (
cycloNTSFTPRL) retains significant activity on the pheromonotropic HevPBANR receptor from the tobacco budworm
Heliothis virescens expressed in CHO-K1 cells. Previous studies indicate that this rigid, cyclic analog adopts a type I β-turn with a
transPro over residues TPRL within the core PK/PBAN region. An analog containing an (
E)-alkene,
trans-Pro mimetic motif was synthesized, and upon evaluation on the HevPBANR receptor found to have an EC
50 value that is not statistically different from a parent C-terminal PK/PBAN hexapeptide sequence. The results, in aggregate, provide strong evidence for the orientation of Pro and the core conformation of PK/PBAN neuropeptides during interaction with the expressed PBAN receptor. The work further identifies a novel scaffold with which to design mimetic PBAN analogs as potential leads in the development of environmentally favorable pest management agents capable of disrupting PK/PBAN-regulated pheromone signaling systems.
The insect kinins are potent diuretic peptides that preferentially form a cis-Pro, type VI β-turn. An insect kinin analog containing (2S,4S)-4-aminopyroglutamate, a novel cis-peptide bond, type VI ...β-turn motif, demonstrates significant activity in the physiological range in a cricket diuretic assay. This is the first instance of a 4-aminopyroglutamate analog of a peptide with a preference for a type VI turn that demonstrates significant bioactivity. The results provide further confirmatory evidence for the active conformation of the insect kinins, and a new scaffold with which to design biostable, peptidomimetic analogs capable of disrupting critical insect kinin-regulated processes in insects.
In the present study we describe the synthesis and some pharmacological properties of eight new analogues of bradykinin (BK). Two peptides were designed by substitution of position 7 or 8 of the ...known D-Arg(0),Hyp(3),Thi(5,8),D-Phe(7)BK antagonist (Stewart's antagonist) with L-pipecolic acid (L-Pip). The next two analogues were obtained by replacement of the d-Phe residue in position 7 of the Stewart's peptide with L-beta(2)-isoproline (L-beta(2)-iPro) or L-beta(3)-homoproline (L-beta(3)-hPro). The four analogues mentioned above were also prepared in N-acylated form with 1-adamantaneacetic acid (Aaa). Biological activity of the compounds was assessed by isolated rat uterus and rat blood pressure tests. Our results showed that L-Pip in position 7 slightly increased antagonistic potency in the blood pressure test, but it turned the analogue into an agonist in the rat uterus test. Replacement of Thi by L-Pip in position 8 also enhanced antagonism in the rat pressure test but preserved the antagonism in the rat uterus test. L-beta(2)-iPro or L-beta(3)-hPro in position 7 decreased the potencies in both tests. We also demonstrated that acylation of the N-terminus did not increase, as was claimed previously, the antagonistic potencies of the resulting peptides. The results thus support the hypothesis about the existence of different types of BK receptors in the rat uterus and blood vessels. Our studies provide new information about the structure-activity relationship of BK antagonists which may help in designing more potent BK receptor blockers.
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