The insect kinins are multifunctional neuropeptides found in a variety of arthropod species, including the pea aphid
Acyrthosiphon pisum (Hemiptera: Aphidae). A series of biostable insect kinin ...analogs based on the shared C-terminal pentapeptide core region were fed in solutions of artificial diet to the pea aphid over a period of 3 days and evaluated for antifeedant and aphicidal activity. The analogs contained either α,α-disubstituted or β-amino acids in key positions to enhance resistance to tissue-bound peptidases and retain activity in a number of insect kinin bioassays and/or on expressed receptors. Three of the biostable analogs demonstrated antifeedant activity, with a marked reduction in honeydew formation observed after 1 day, and very high mortality. In contrast, an unmodified, parent insect kinin and two other analogs containing some of the same structural components that promote biostability are inactive. The most active analog, double Aib analog
K-Aib-1 (AibFFAibWGa), featured aphicidal activity calculated at an LC
50 of 0.063
nmol/μl (0.048
μg/μl) and an LT
50 of 1.68 days, matching the potency of some commercially available aphicides. The mechanism of this activity has yet to be established. The aphicidal activity of the biostable insect kinin analogs may result from different potential mechanisms as disruption of digestive processes by interfering with gut motility patterns, digestive enzyme release, and/or with fluid cycling in the gut, and also nutrient transport across the gut itself; all processes shown to be regulated by the insect kinins in other insects. However the mechanism(s) is(are) not yet known. The active insect kinin analogs represent potential leads in the development of selective, environmentally friendly pest aphid control agents.
Abstract The multifunctional arthropod ‘insect kinins’ share the evolutionarily conserved C-terminal pentapeptide motif Phe-X1 -X2 -Trp-Gly-NH2 , where X1 = His, Asn, Ser, or Tyr and X2 = Ser, Pro, ...or Ala. Insect kinins regulate diuresis in many species of insects. Compounds with similar biological activity could be exploited for the control of arthropod pest populations such as the mosquito Aedes aegypti (L.) and the southern cattle tick Rhipicephalus ( Boophilus) microplus (Canestrini), vectors of human and animal pathogens, respectively. Insect kinins, however, are susceptible to fast enzymatic degradation by endogenous peptidases that severely limit their use as tools for pest control or for endocrinological studies. To enhance resistance to peptidases, analogs of the insect kinins incorporating bulky α,α-disubstituted amino acids in positions adjacent to both primary and secondary peptidase hydrolysis sites were synthesized. In comparison with a control insect kinin, several of these analogs are highly stable to hydrolysis by degradative enzymes ANCE, neprilysin and Leucine aminopeptidase. Six analogs were evaluated by calcium bioluminescence assay on recombinant receptors from mosquito and tick. Four of these analogs either matched or exceeded the potency of the control kinin peptide agonist. One of these was about 5-fold more potent than the control agonist on the tick receptor. This analog was 8-fold more potent than the control agonist on the mosquito receptor, and twice more potent than the endogenous Aedes kinin-II. The analog also demonstrated potent activity in an in vitro Aedes Malpighian tubule fluid secretion assay. Similar comparisons of analog potency cannot be made to tick kinins because no endogenous kinin has yet been identified. These potent, biostable analogs represent ideal new tools for endocrinologists studying arthropod kinin-regulated processes in vivo , particularly for ticks in which their role remains to be established.
Diapause hormone (DH) is a peptide well known to induce embryonic diapause in the commercial silkmoth
Bombyx mori. More recently, this same neuropeptide was reported to break diapause in pupae of the ...agriculturally important
Heliothis/Helicoverpa complex. In this study we examine the efficacy and potency of a select group of structural analogs of the native hormone in
Helicoverpa zea and report the structures of several analogs that are considerably more potent than DH in breaking diapause. Among the most potent analogs (PK-Etz, PK-2Abf, 901) were those with structural components that enhance resistance to peptidases that degrade and inactivate the native peptide
in vivo, which may account, at least in part, for the observed increase in potency for these analogs. Analog 901 was previously demonstrated to both enhance biostablility and bioavailability properties in adult heliothines and thus may be a potential candidate for topical application as a diapause-terminating agent. The significant activity observed for two restricted conformation analogs is consistent with an active conformation for diapause hormone that features a
transPro within a type I β-turn in the C-terminal region. DH is also known to successfully break diapause only within a fairly narrow temperature range. While DH is effective at 21
°C, it is not effective at 18
°C. Likewise, the analogs were effective at 21
°C but not at 18
°C. By contrast, 20-hydroxyecdysone, a steroid hormone that is also capable of breaking diapause is effective at both temperatures, thus suggesting that DH and the ecdysteroids act through different mechanisms to terminate diapause.
A linear pyrokinin (PK)/pheromone biosynthesis activating neuropeptide (PBAN) antagonist lead (RYFdFPRLa) was structurally modified to impart amphiphilic properties to enhance its ability to ...transmigrate the hydrophobic cuticle of noctuid moth species and yet retain aqueous solubility in the hemolymph to reach target PK/PBAN receptors within the internal insect environment. The resulting novel PK/PBAN analog, Hex-Suc-AdFPRLa (PPK-AA), was synthesized and evaluated as an antagonist in a pheromonotropic assay in
Heliothis peltigera against 4 natural PK/PBAN peptide elicitors (PBAN; pheromonotropin, PT; myotropin, MT; leucopyrokinin, LPK) and in a melanotropic assay in
Spodoptera littoralis against 3 natural PK/PBAN peptide elicitors (PBAN, PT, LPK). The analog proved to be a potent and efficacious inhibitor of sex pheromone biosynthesis elicited by PBAN (84% at 100
pmol) and PT (54% at 100
pmol), but not by MT and LPK. PPK-AA is a selective pure antagonist (i.e., does not exhibit any agonistic activity) as it failed to inhibit melanization elicited by any of the natural PK/PBAN peptides. The analog was shown to transmigrate isolated cuticle dissected from adult female
Heliothis virescens moths to a high extent of 25–30% (130–150
pmol), representing physiologically significant quantities. PPK-AA represents a significant addition to the arsenal of tools available to arthropod endocrinologists studying the endogenous mechanisms of PK/PBAN regulated processes, and a prototype for the development of environmentally friendly pest management agents capable of disrupting the critical process of reproduction.
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 ability of linear β-amino acid substituted peptides (PK-βA-1: Ac-YFTβ
3PRLa; PK-βA-2: Ac-Yβ
3homoFTPRLa; PK-βA-3: Ac-Yβ
3FTPRLa; PK-βA-4: Ac-β
3FFTβ
3PRLa) and unsubstituted analogs (Ac-YFTPRLa ...and YFTPRLa) of the pyrokinin(PK)/pheromone biosynthesis-activating neuropeptide (PBAN) family to penetrate the insect cuticle and exert biological activity (i.e., stimulate sex pheromone biosynthesis), was tested by topical application on
Heliothis peltigera moths. The present results clearly indicate that small linear synthetic peptides can penetrate the cuticle very efficiently by contact application and activate their target organ. The time responses of the peptides applied in DDW and DMSO were tested and the activities of topically applied and injected peptides were compared. The results clearly indicate that PK-βA-4 and PK-βA-3 exhibited high bioavailability (ability to penetrate through the cuticle and exertion of bioactivity) with the latter showing longer persistence in both solvents than any other analog in the study; indicative that incorporation of a β-amino acid at the Phe
2 position can enhance longevity in topical PK/PBAN analogs. PK-βA-4 was significantly more active in DMSO than in DDW, and significantly more active than the parent peptide LPK in DMSO. PK-βA-1 and PK-βA-2 exhibited negligible activity. Interestingly, Ac-YFTPRLa was highly potent in both solvents; its activity in DDW did not differ from that of PK-βA-4 and PK-βA-3, and was higher than that of LPK. Even the unacylated peptide YFTPRLa was active in both solvents, at a similar level to LPK. Topically applied PK-βA-4 and Ac-YFTPRLa exhibited significantly higher activity than the injected peptides. PK-βA-3 and YFTPRLa were equally potent in both routes of administration.
The multifunctional arthropod ‘insect kinins’ share the evolutionarily conserved C-terminal pentapeptide motif Phe-X
1-X
2-Trp-Gly-NH
2, where X
1
=
His, Asn, Ser, or Tyr and X
2
=
Ser, Pro, or Ala. ...Eight different analogs of the insect kinin C-terminal pentapeptide active core in which the critical residues Phe
1, Pro
3 and Trp
4 are replaced with β
3-amino acid and/or their β
2-amino acid counterparts were evaluated on recombinant insect kinin receptors from the southern cattle tick,
Boophilus microplus (Canestrini) and the dengue vector, the mosquito
Aedes aegypti (L.). A number of these analogs previously demonstrated enhanced resistance to degradation by peptidases. Single-replacement analog β
2Trp
4 and double-replacement analog β
3Phe
2, β
3Pro
3 of the insect kinins proved to be selective agonists for the tick receptor, whereas single-replacement analog β
3Pro
3 and double-replacement analog β
3Phe, β
3Pro
3 were strong agonists on both mosquito and tick receptors. These biostable analogs represent new tools for arthropod endocrinologists and potential leads in the development of selective, environmentally friendly arthropod pest control agents capable of disrupting insect kinin-regulated processes.
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