Biomedical subjects

J M Conlon

Publications and source records attributed to J M Conlon.

At least 19 recordsLinked to original sources

PubMed2001

Antimicrobial peptides isolated from skin secretions of the diploid frog, Xenopus tropicalis (Pipidae).

Seven peptides (XT-1-XT-7) with antimicrobial activity were isolated from norepinephrine-stimulated skin secretions of the diploid clawed frog, Xenopus tropicalis. Structural characterization of the peptides demonstrated that amino acid sequence similarity to antimicrobial peptides previously isolated from Xenopus laevis was low, suggesting that the species are not closely related phylogenetically. Peptides XT-5 and XT-3 are probably the orthologs of X. laevis peptide glycine-leucine amide (PGL(a)) and the N-terminal spacer region of prolevitide, respectively. XT-1, XT-6 and XT-7 show limited structural similarity to the spacer region of X. laevis procaeruleins and the paralogs XT-2 and XT-4 are similar to corresponding regions of proxenopsin. Orthologs of the magainins were not identified. The C-terminally alpha-amidated peptide XT-7 (GLLGPLLKIAAKVGSNLL.NH2) showed the lowest minimum inhibitory concentrations against reference microorganisms (Staphylococcus aureus 5 microM, Escherichia coli 5 microM, and Candida albicans 40 microM) and was also active against clinical isolates of methicillin-resistant S. aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus, Streptococcus group C, Shigella sonnei, Pseudomonas aeruginosa and Enterobacter cloacae. The peptide was, however, hemolytic against human erythrocytes (50% lysis at 70 microM). Circular dichroism studies showed that XT-7 has a random structure in aqueous solution, pH 7.0 but adopts an alpha-helical conformation in the presence of 50% trifluoroethanol. Decreasing the cationicity of XT-7 either by replacement of the C-terminal CONH2 group by COOH or by deletion of the Lys(8) residue produced analogs with greatly (>10-fold) decreased antimicrobial potencies.

Amino Acid Sequence↗

PubMed2001

Pseudin-2: an antimicrobial peptide with low hemolytic activity from the skin of the paradoxical frog.

Four structurally related peptides (pseudins 1-4) with antimicrobial activity were isolated from an extract of the skin of the paradoxical frog Pseudis paradoxa (Pseudidae). Pseudin-2 (GLNALKKVFQGIHEAIKLINNHVQ) was the most abundant peptide (22 nmol/g tissue) and also the most potent (minimum inhibitory concentrations, MIC = 2.5 microM against Escherichia coli, 80 microM against Staphylococcus aureus, and 130 microM against Candida albicans). The concentration of pseudin-2 producing 50% hemolysis of human erythrocytes was >300 microM. Circular dichroism studies showed that the pseudins belong to the class of cationic, amphipathic alpha-helical antimicrobial peptides but their amino acid sequences are not similar to any previously characterized peptides from frog skin. The pseudins do, however, show sequence similarity with a region at the C-terminus of DEFT, a death effector domain-containing protein expressed in mammalian testicular germ cells that is involved in the regulation of apoptosis.

Amino Acid Sequence↗

PubMed2001

Pituitary adenylate cyclase-activating polypeptide and its receptors in amphibians.

Pituitary adenylate cyclase-activating polypeptide (PACAP), a novel peptide of the secretin/glucagon/vasoactive intestinal polypeptide superfamily, has been initially characterized in mammals in 1989 and, only 2 years later, its counterpart has been isolated in amphibians. A number of studies conducted in the frog Rana ridibunda have demonstrated that PACAP is widely distributed in the central nervous system (particularly in the hypothalamus and the median eminence) and in peripheral organs including the adrenal gland. The cDNAs encoding the PACAP precursor and 3 types of PACAP receptors have been cloned in amphibians and their distribution has been determined by in situ hybridization histochemistry. Ontogenetic studies have revealed that PACAP is expressed early in the brain of tadpoles, soon after hatching. In the frog Rana ridibunda, PACAP exerts a large array of biological effects in the brain, pituitary, adrenal gland, and ovary, suggesting that, in amphibians as in mammals, PACAP may act as neurotrophic factor, a neurotransmitter and a neurohormone.

Adrenal Glands↗

PubMed2001

Amphibian glucagon family peptides: potent metabolic regulators in fish hepatocytes.

Peptides analogous to glucagon-like peptide-1 (GLP-1) have been isolated from amphibian pancreas and intestine, and their amino acid sequences and cDNA structures elucidated. Just like their mammalian counterpart, these peptides are potent insulinotropins in mammalian pancreatic cells. We show here that these peptides also exert strong glycogenolytic actions when applied to dispersed fish hepatocytes. We compared the potencies of three synthetic GLP-1s from Xenopus laevis and two native GLP-1s from Bufo marinus in the activation of glycogenolysis in the hepatocytes of a marine rockfish (Sebastes caurinus) and two freshwater catfish (Ameiurus nebulosus and A. melas), and demonstrated their effectiveness in increasing the degree of phosphorylation of glycogen phosphorylase. We also compared the glycogenolytic potency of the peptides with those of human GLP-1 and glucagons from human and B. marinus. Sensitivity to these peptides is species-specific, with the rockfish responding at lower concentrations to GLP-1s and the two catfish reacting better to glucagons. However, the relative potency of the amphibian GLP-1s and glucagons is similar in the three species. Xenopus GLP-1C (xGLP-1C) is consistently more potent than xGLP-1B, while xGLP-1A displays the smallest activation of glycogenolysis. Similarly, Bufo GLP-1(32)-the peptide with the highest amino acid sequence identity to xGLP-1C-always shows a higher potency than Bufo GLP-1(37), which is closely related to xGLP-1B. The relative hierarchy of these glycogenolytic GLP-1s differs from their ranking as insulinotropins in mammalian beta-cells. In the rockfish system, Bufo glucagon-36, a C-terminally extended glucagon, is more potent than the shorter bovine glucagon and Bufo glucagon-29 in the activation of glycogenolysis; when tested in A. nebulosus hepatocytes, bovine and amphibian glucagons are equipotent. Amphibian GLP-1s and glucagons activate glycogenolysis in fish hepatocytes through increased phosphorylation of glycogen phosphorylase, implying involvement of the adenylyl cyclase/protein kinase A system in signal transduction. We conclude that the broad physiological effectiveness of GLP-1 has been retained throughout vertebrate evolution, and that both insulinotropic activity and glycogenolytic actions belong to the repertoire of GLP-1.

Amino Acid Sequence↗

PubMed2001

The endocrine cells in the gastroenteropancreatic system of the bowfin, Amia calva L.: an immunohistochemical, ultrastructural, and immunocytochemical analysis.

The gastroenteropancreatic (GEP) endocrine system of bowfin (Amia calva) was described using light and electron microscopy and immunological methods. The islet organ (endocrine pancreas) consists of diffusely scattered, mostly small islets and isolated patches of cells among and within the exocrine acini. The islets are composed of abundant, centrally located B cells immunoreactive to bovine and lamprey insulin antisera and D cells showing a widespread distribution and specificity to somatostatin antibodies. A and F cells are present at the very periphery of the islets and are immunoreactive with antisera against glucagon (and glucagon-like peptide) and several peptides of the pancreatic polypeptide (PP)-family, respectively. The peptides of the two families usually collocates within the same peripheral islet cells and are the most common immunoreactive peptides present in the extra-islet tissue. Immunocytochemistry and fine structural observations characterised the granule morphology for B and D cells and identified two cell types with granules immunoreactive to glucagon antisera. These two putative A cells had similar granules, which were distinct from either B or D cells, but one of the cells had rod-shaped cytoplasmic inclusions within cisternae of what appeared to be rough endoplasmic reticulum. The inclusions were not immunoreactive to either insulin or glucagon antisera. Only small numbers of cells in the stomach and intestine immunoreacted to antisera against somatostatin, glucagon, and PP-family peptides. The paucity of these cells was reflected in the low concentrations of these peptides in intestinal extracts. The GEP system of bowfin is not unlike that of other actinopterygian fishes, but there are some marked differences that may reflect the antiquity of this system and/or may be a consequence of the ontogeny of this system in this species.

Animals↗

PubMed2001

Effects of trout endothelin on the motility of gastrointestinal smooth muscle from the trout and rat.

Trout endothelin (ET), previously isolated from the kidney of the rainbow trout Oncorhynchus mykiss, contains four amino acid substitutions at residues 4-7 compared with rat ET-1. Trout ET produced sustained and concentration-dependent contractions of strips of longitudinal smooth muscle from trout stomach (pD(2) = 7.52 +/- 0.06) and proximal small intestine (pD(2) = 7.80 +/- 0.10) and from rat fundus (pD(2) = 7.78 +/- 0.14). Rat ET-1 was equipotent with trout ET for contraction of rat fundus and 2- to 3-fold more potent for contraction of trout gastrointestinal tissues. In contrast, rat ET-1 was 10- to 20-fold more potent than trout ET in constricting isolated rings of vascular tissue from trout efferent branchial artery and cardinal vein and from rat aorta (Y. Wang et al., 1999, Am. J. Physiol. 277, R1605-R1611). It is known that the contractile effects of ET-1 on rat fundus are mediated through the ET(B) receptor and effects on the rat aorta are mediated through the ET(A) receptor. We propose, therefore, that trout gastrointestinal tissues express an ET(B)-type receptor that differentiates poorly between trout ET and rat ET-1, whereas trout vascular tissues express an ET(A)-type receptor that is preferentially activated by rat ET-1. The rat ET-1-induced contractions of the trout gastrointestinal tissues are in part indirect, involving a serotoninergic neuronal pathway in the intestine and a noncholinergic, nonserotoninergic pathway in the stomach.

Animals↗

PubMed2001

Anomalous rates of evolution of pancreatic polypeptide and peptide tyrosine-tyrosine (PYY) in a tetraploid frog, Xenopus laevis (Anura:Pipidae).

The South African clawed frog Xenopus laevis is believed to have arisen as a result of a tetraploidization event occurring approximately 30 million years ago. Two molecular forms of pancreatic polypeptide (PP) have been isolated from an extract of the pancreas of this species and two molecular forms of peptide tyrosine-tyrosine (PYY) from the intestine. Despite the fact that the amino acid sequence of PP has, in general, been very poorly conserved during the evolution of tetrapods (only Pro(5), Pro(8), Gly(9), Ala(12), Tyr(27), Arg(33) and Arg(35) are invariant among species studied so far), the two Xenopus PPs differ by only a single amino acid substition (Asp(22)-->Glu). In contrast the two molecular forms of PYY differ by six amino acid substitutions (Glu(15)-->Gln, Thr(18)-->Ala, Leu(21)-->Met, Ile(22)-->Thr, Ile(28)-->Val, Val(31)-->Ile). The data imply that strong evolutionary pressure is acting to conserve the functional domain in both genes encoding PP and so suggest that PP may have an important physiological role in amphibians (although the nature of this role has yet to be determined). The more rapid mutation of the functional domain in the genes encoding PYY, a peptide whose amino acid sequence has been quite well conserved in tetrapods and whose physiological significance is well established, suggests that one of the PYY genes may be evolving towards a new function or towards becoming a pseudogene.

Amino Acid Sequence↗

PubMed2001

Characterization and distribution of neuropeptide Y in the brain of a caecilian amphibian.

Neuropeptide Y (NPY) from the brain of an amphibian from the order Gymnophiona (the caecilian, Typhlonectes natans) was characterized. We cloned a 790 base pair cDNA encoding the caecilian NPY precursor. The open reading frame consisted of 291 bases, indicating an NPY precursor of 97 amino acids. Both deduced and isolated NPY primary structures were Tyr-Pro-Ser-Lys-Pro-Asp-Asn-Pro-Gly-Glu(10)-Asp-Ala-Pro-Ala-Glu-Asp-Met-Ala-Lys-Tyr(20)-Tyr-Ser-Ala-Leu-Arg-His-Tyr-Ile-Asn-Leu(30)-Ile-Thr-Arg-Gln-Arg-Tyr. NH2. In caecilian brain, we observed NPY immunoreactive cells within the medial pallium, basal forebrain, preoptic area, midbrain tegmentum and trigeminal nucleus. The prevalence of preoptic and hypothalamic terminal field staining supports the hypothesis that NPY controls pituitary function in this caecilian.

Amino Acid Sequence↗

PubMed2001

Effects of cod bradykinin and its analogs on vascular and intestinal smooth muscle of the Atlantic cod, Gadus morhua.

The effects of [Arg(0),Trp(5),Leu(8)]-BK (cod [Arg(0)]BK) on vascular preparations from branches of the cod celiac artery and on longitudinal smooth muscle preparations from the cod intestine were investigated. Cod [Arg(0)]BK (3 x 10(-8) M) caused a relaxation of the celiac artery precontracted with adrenaline. The relaxation was abolished by the cyclooxygenase inhibitor indomethacin, suggesting that the effect is mediated through the release of prostaglandins, but there was no evidence for the involvement of leukotrienes or nitric oxide in the response. In the intestinal preparations, cod [Arg(0)]BK produced concentration-dependent contractions (pD(2) = 8.28 +/- 0.16). Experiments with N-terminally and C-terminally truncated analogs and with alanine-substituted analogs of cod [Arg(0)]BK demonstrate that the central amino acid Gly(4) and the C-terminal amino acids Leu(8) and Arg(9) are the most important in determining the conformation of the peptide that interacts with the receptor. The results indicate that the ligand binding properties of the cod BK receptor are considerably different from the receptor present in trout tissues and may resemble those of the mammalian B(2) receptor more closely.

Alanine↗

PubMed2001

Evolution of the insulin molecule: insights into structure-activity and phylogenetic relationships.

The conformation of insulin in the crystalline state has been known for more than 30 years but there remains uncertainty regarding the biologically active conformation and the structural features that constitute the receptor-binding domain. The primary structure of insulin has been determined for at least 100 vertebrate species. In addition to the invariant cysteines, only ten amino acids (GlyA1, IleA2, ValA3, TyrA19, LeuB6, GlyB8, LeuB11, ValB12, GlyB23 and PheB24) have been fully conserved during vertebrate evolution. This observation supports the hypothesis derived from alanine-scanning mutagenesis studies that five of these invariant residues (IleA2, ValA3, TyrA19, GlyB23, and Phe24) interact directly with the receptor and five additional conserved residues (LeuB6, GlyB8, LeuB11, GluB13 and PheB25) are important in maintaining the receptor-binding conformation. With the exception of the hagfish, only conservative substitutions are found at B13 (Glu --> Asp) and B25(Phe --> Tyr). In contrast, amino acid residues that were also considered to be important in receptor binding based upon the crystal structure of insulin (GluA4, GlnA5, AsnA21, TyrB16, TyrB26) have been much less well conserved and are probably not components of the receptor-binding domain. The hypothesis that LeuA13 and LeuB17 form part of a second receptor-binding site in the insulin molecule finds some support in terms of their conservation during vertebrate evolution, although the site is probably absent in some hystricomorph insulins. In general, the amino acid sequences of insulins are not useful in cladistic analyses especially when evolutionary distant taxa are compared but, among related species in a particular order or family, the presence of unusual structural features in the insulin molecule may permit a meaningful phylogenetic inference. For example, analysis of insulin sequences supports monophyletic status for Dipnoi, Elasmobranchii, Holocephali and Petromyzontiformes.

Alanine↗

PubMed2001

The structure of Mordacia mordax insulin supports the monophyly of the Petromyzontiformes and an ancient divergence of Mordaciidae and Geotriidae.

The phylogenetic relationships between the two southern hemisphere lamprey families (Geotriidae and Mordaciidae) and their northern hemisphere counterparts (Petromyzontidae) are unresolved. Insulin was isolated from an extract of islet-containing intestinal tissue from ammocoetes of the Australian lamprey, Mordacia mordax. Its primary structure was established as A-chain: GIVEQCCHRK10CSIYDMENYC20N and B-chain: SALMGTGGTH10LCGSHLVEAL20YVVCGQRGFF30 YTP[SKG]. Although the residues in parentheses are only tentatively assigned, mass spectrometry supports the proposed sequence and demonstrates that Mordacia proinsulin, unlike proinsulin from Geotria australis, is fully processed to mature insulin. Insulins from M. mordax and G. australis and from the northern hemisphere lampreys Petromyzon marinus and Lampetra fluviatilis share a pentapeptide extension to N-terminus of the B-chain (Ser-Ala-Leu-Xaa-Gly) that has never been found in the insulins of any other vertebrate class. This observation provides support for the claim that the Petromyzontiformes constitute a monophyletic group. M. mordax insulin differs from that of G. australis by 18 amino acid residues but by only four residues from the common sequence of P. marinus and L. fluviatilis insulin. These data are consistent with the view that Geotriidae and Mordaciidae have been separated for a long period and suggest that G. australis insulin has undergone an accelerated rate of molecular evolution.

Amino Acid Sequence↗

PubMed2001

Antimicrobial peptides from the skin of the Japanese mountain brown frog, Rana ornativentris.

Six peptides with antimicrobial activity were isolated from an extract of freeze-dried skin of the Japanese mountain brown frog Rana ornativentris. Two structurally related peptides (brevinin-20a GLFNVFKGALKTAGKHVAGSLLNQLKCKVSGGC, 11 nmol/g dried tissue, and brevinin-20b GIFNVFKGALKTAGKHVAGSLLNQLKCKVSGEC, 170 nmol/g) belong to the brevinin-2 family, previously identified in Asian and European, but not North American, Ranid frogs. Four peptides (temporin-10a FLPLLASLFSRLL.NH2, 13 nmol/g; temporin-10b FLPLIGKILGTI L.NH2, 350 nmol/g; temporin-10c FLPLLASLFSRLF.NH2, 14 nmol/g; and temporin-10d FLPLLASLFSGLF.NH2, 8 nmol/g) are members of the temporin family first identified in the European common frog Rana temporaria but also found in the skins of North American Ranids. The brevinin-2 peptides showed broad-spectrum activity against the gram-positive bacterium, Staphylococcus aureus, the gram-negative bacterium, Escherichia coli and the yeast Candida albicans, whereas the temporins showed potent activity only against S. aureus. The brevinins and temporins belong to the class of cationic antimicrobial peptides that adopt an amphipathic alpha-helical conformation but it is significant that temporin-10d, which lacks a basic amino acid residue, is still active against S. aureus (minimum inhibitory concentration=13 microM compared with 2 microM for temporin-10a). This suggests that strong electrostatic interaction between the peptide and the negatively charged phospholipids of the cell membrane is not an absolute prerequisite for antimicrobial activity.

Amino Acid Sequence↗

PubMed2001

A neuropeptide Y receptor Y1-subfamily gene from an agnathan, the European river lamprey. A potential ancestral gene.

We report here the isolation and functional expression of a neuropeptide Y (NPY) receptor from the river lamprey, Lampetra fluviatilis. The receptor displays approximately 50% amino-acid sequence identity to all previously cloned Y1-subfamily receptors including Y1, Y4, and y6 and the teleost subtypes Ya, Yb and Yc. Phylogenetic analyses point to a closer relationship with Y4 and Ya/b/c suggesting that the lamprey receptor could possibly represent a pro-orthologue of some or all of those gnathostome receptors. Our results support the notion that the Y1 subfamily increased in number by genome or large-scale chromosome duplications, one of which may have taken place prior to the divergence of lampreys and gnathostomes whereas the second duplication probably occurred in the gnathostome lineage after this split. Functional expression of the lamprey receptor in a cell line facilitated specific binding of the three endogenous lamprey peptides NPY, peptide YY and peptide MY with picomolar affinities. Binding studies with a large panel of NPY analogues revealed indiscriminate binding properties similar to those of another nonselective Y1-subfamily receptor, zebrafish Ya. RT-PCR detected receptor mRNA in the central nervous system as well as in several peripheral organs suggesting diverse functions. This lamprey receptor is evolutionarily the most distant NPY receptor that clearly belongs to the Y1 subfamily as defined in mammals, which shows that subtypes Y2 and Y5 arose even earlier in evolution.

Amino Acid Sequence↗

PubMed2001

Gastrin-releasing peptides from Xenopus laevis: purification, characterization, and myotropic activity.

Two molecular forms of gastrin-releasing peptide (GRP) were isolated from an extract of the intestine of the tetraploid frog Xenopus laevis. The primary structure of GRP-1 (APTSQQHTEQ(10)LSRSNINTRG(20) SHWAVGHLM.NH(2)) differs from that of GRP-2 by a single amino acid substitution (Asn(15)--> Thr(15)). GRP-(20-29) peptide (neuromedin C) was also isolated from the extract. Synthetic GRP-1 produced concentration-dependent contractions of longitudinal smooth muscle strips from Xenopus cardiac stomach (pD(2) = 8.93 +/- 0.32; n = 6). The responses were unaffected by tetrodotoxin, atropine, and methysergide, indicating a direct action of the peptide on smooth muscle cells. GRP-1 elicited concentration-dependent relaxations of precontracted (5 microM carbachol) circular smooth muscle strips from the same region (pD(2) = 8.96 +/- 0.21; n = 8). The responses were significantly (P < 0.05) attenuated (71 +/- 24% decrease in maximum response; n = 6) by indomethacin, indicating mediation, at least in part, by prostanoids. Despite the fact that Xenopus GRP-1 differs from pig GRP at 15 amino acid sites, both peptides are equipotent and equally effective for both contractile and relaxant responses, demonstrating that selective evolutionary pressure has acted to conserve the functional COOH-terminal domain in the peptide. The data suggest a physiologically important role for GRP in the regulation of gastric motility in X. laevis.

Amino Acid Sequence↗

PubMed2001

Antidipsogenic effects of eel bradykinins in the eel Anguilla japonica.

A peptide with bradykinin (BK)-like immunoreactivity was isolated from an incubate of heat-denatured eel plasma with porcine pancreatic kallikrein. The purified peptide had the following amino acid sequence: Arg-Arg-Pro-Pro-Gly-Ser-Trp-Pro-Leu-Arg. This decapeptide, named eel [Arg(0)]BK, was identical to two previously identified BK homologs from cod and trout. High conservation of the BK sequence among distant teleost species suggests an important function in this vertebrate group. Bolus intra-arterial injections of eel [Arg(0)]BK, BK, and [Arg(0)]-des-Arg(9)-BK (1-10 nmol/kg) caused significant (P < 0.05) inhibition of drinking in seawater-adapted eels. The potency of the inhibition was ranked in the following order: [Arg(0)]BK > [Arg(0)]-des-Arg(9)-BK = BK. The BK peptides also produced an immediate, transient increase followed by a sustained increase in arterial blood pressure and an initial decrease followed by an increase in heart rate. Strong tachyphylaxis occurred for the cardiovascular effect but not for the antidipsogenic effect. The order of the potency of the cardiovascular actions, [Arg(0)]BK > BK > [Arg(0)]-des-Arg(9)-BK, was different from that of the antidipsogenic action. Slow infusions of eel [Arg(0)]BK in the dose range 1-1,000 pmol x kg(-1) x min(-1) produced concentration-dependent inhibition of drinking without changes in arterial pressure, plasma osmolality, and hematocrit. At the infusion rate of >100 pmol x kg(-1) x min(-1), plasma concentrations of angiotensin II, a potent dipsogenic hormone in eels, increased, suggesting an interaction of the kallikrein-kinin and renin-angiotensin systems. In mammals, BK is dipsogenic and vasodepressor, so that our data demonstrate opposite effects on fluid and cardiovascular regulation of BK in the eel and suggest a new physiological role for the kallikrein-kinin system in teleost fish.

Angiotensin II↗

PubMed2000

A protease inhibitor of the Kunitz family from skin secretions of the tomato frog, Dyscophus guineti (Microhylidae).

Norepinephrine-stimulated skin secretions of the tomato frog, Dyscophus guineti, contained a trypsin inhibitor whose primary structure was established as: SPAEVCF LPK(10) ESGLCRARAL(20) RYYYDRGDGK(30) CEEFIYGGCG(40) GNGNNY KSLL(50) TCKISCE. This amino acid sequence identifies the peptide as a member of the Kunitz/bovine pancreatic trypsin inhibitor (BPTI) family and demonstrates that selective evolutionary pressure has acted to conserve those domains in the molecule (corresponding to positions 12-18 and 34-39 in BPTI) that interact with trypsin. Extracellular proteases produced by pathogenic microorganisms play important roles in facilitating invasion of the host and broad spectrum antimicrobial activity of BPTI has been described. Cationic, amphipathic alpha-helical antimicrobial peptides of the magainin type, important in the defense strategy of several species of frog, were not detected in the skin secretions. We speculate, therefore, that synthesis of a proteinase inhibitor in the skin of the tomato frog may be a component of an alternative strategy of this animal to defend itself against microorganisms.

Adrenergic alpha-Agonists↗

PubMed2000

Multiple antimicrobial peptides and peptides related to bradykinin and neuromedin N isolated from skin secretions of the pickerel frog, Rana palustris.

The skin secretions of the North American pickerel frog Rana palustris are toxic to both microorganisms and predators. A total of 22 peptides with differential growth-inhibitory activity towards bacteria and yeast were isolated from the electrostimulated secretions of R. palustris skin and were characterized structurally. Thirteen of the antimicrobial peptides belong to five of the known families previously identified in the skins of other species of Ranid frogs: brevinin-1 (3 peptides), esculentin-1 (2 peptides), esculentin-2 (1 peptide), ranatuerin-2 (6 peptides), and temporin (1 peptide). Nine peptides show little structural similarity towards other known antimicrobial peptides and so are classified in new families: palustrin-1 (4 peptides) with 27-28 amino acid residues and a cystine-bridged heptapeptide ring; palustrin-2 (3 peptides) with 31 amino acids and a cyclic heptapeptide region and palustrin-3 (2 peptides) with 48 amino acids and a cyclic hexapeptide region. Peptides belonging to the esculentin-1, esculentin-2 and palustrin-3 families are the most potent (minimal inhibitory concentrations approximately 1 microM against Escherichia coli) whereas peptides of the brevinin-1 and esculentin-2 families show the broadest spectrum of activity. As well as bradykinin that is identical to the human peptide, a further 4 peptides structurally related to [Leu(8)]bradykinin and two peptides related to neuromedin-N (the hexapeptide KKPYIL and a larger, cystine-containing form HLRRCGKKPYILMACS) were purified from the skin secretions.

Amino Acid Sequence↗

PubMed2000

Induction of synthesis of an antimicrobial peptide in the skin of the freeze-tolerant frog, Rana sylvatica, in response to environmental stimuli.

An extract of skin taken from specimens of the freeze-tolerant wood frog, Rana sylvatica, that were collected from cold (<7 degrees C) ponds and maintained at 5 degrees C lacked detectable antimicrobial activity. In contrast, an extract of skin taken from specimens maintained at 30 degrees C for 3 weeks under laboratory conditions contained a high concentration (approximately 4 nmol/g) of a single antimicrobial peptide of the brevinin-1 family (FLPVVAGLAAKVLPSIICAVTKKC). The peptide inhibited growth of Escherichia coli (minimum inhibitory concentration 45 microM) and Staphylococcus aureus (minimum inhibitory concentration 7 microM). The data suggest that synthesis of the peptide is induced when the animal is in an environment that promotes the growth of microorganisms consistent with a role in the animal's defense strategy.

Adaptation, Physiological↗