Two families of antimicrobial peptides with multiple functions from skin of rufous-spotted torrent frog, Amolops loloensis

There are around 27 species of Amolops amphibian distributed in South-east of Asia. Seven antimicrobial peptides (AMPs) belonging to two different families were purified from skin of rufous-spotted torrent frog, Amolops loloensis, and designated brevinins

Two antimicrobial peptides from skin secretions of Rana grahami

Two antimicrobial peptides manifested a broad spectrum of anti microbial activity against various microorganisms have been isolated from skin secretions of Rana grahami. These antimicrobial peptides were named grahamin 1 and grahamin 2. Their primary Stru

A novel algesic peptide derived from skin secretions of the frog Amolops loloensis

Several algesic agents including bradykinin and tachykinin have been identified from skin secretions of amphibians. They may act in defensive roles against aggressors. In this study, a novel peptide named Amolos with an amino acid sequence of FLPIVGAKL an

Bombinakinin M gene associated peptide, a novel bioactive peptide from skin secretions of the toad Bombina maxima

A novel 28-amino acid peptide, termed bombinakinin-GAP, was purified and characterized from skin secretions of the toad Bombina maxima. Its primary structure was established as DMYEIKQYKTAHGRPPICAPGEQCPIWV-NH2, in which two cysteines form a disulfide bond. A FASTA search of SWISS-PROT databank detected a 32% sequence identity between the sequences of the peptide and a segment of rat cocaine- and amphetamine-regulated transcript (CART). Intracerebroventricular (i.c.v.) administration of the peptide induced a significant decrease in food intake in rats, suggesting that it played a role in the control of feeding by brain. Analysis of its cDNA structure revealed that this peptide is coexpressed with bombinakinin M, a bradykinin-related peptide from the same toad. Bombinakinin-GAP appears to be the first example of a novel class of bioactive peptides from amphibian skin, which may be implicated in feeding behavior. (C) 2003 Elsevier Science Inc. All rights reserved.

A new family of antimicrobial peptides from skin secretions of Rana pleuraden

While conducting experiments to investigate antimicrobial peptides of amphibians living in the Yunnan-Guizhou region of southwest China, a new family of antimicrobial peptides was identified from skin secretions of the Yunnan frog, Rana pleuraden. Members

Isolation and cDNA cloning of cholecystokinin from the skin of Rana nigrovittata

Many neuroendocrine peptides that are distributed in amphibian gastrointestinal tract and central nervous system are also found in amphibian skins, and these peptides are classified into skin-gut-brain triangle peptides, such as bombesins, gastrin-releasi

Cloning of novel bombesin precursor cDNAs from skin of Bombina maxima

Amphibian skin is a rich resource of bioactive peptides like proline-rich bombesin from frog Bombina maxima. A novel cDNA clone encoding a precursor protein that comprises proline-rich bombesin and a novel peptide, designated as bombestatin, was isolated from a skin cDNA library of B. maxima. The predicted primary structure of the novel peptide is WEVLLNVALIRLELLSCRSSKDQDQKESCGMHSW, in which two cysteines form a disulfide bond. A BLAST search of databases did not detect sequences with significant similarity. Bombestatin possesses dose-dependent contractile activity on rat stomach strips. The differences between cDNAs encoding PR-bombesin plus bombestatin and PR-bombesin alone are due to fragment insertions located in 3'-coding region and 3'-untranslational region, respectively. (c) 2005 Elsevier B.V. All rights reserved.

A novel heme-containing protein with anti-HIV-1 activity from skin secretions of Bufo andrewsi

A novel protein, named BAS-AH, was purified and characterized from the skin of the toad Bufo andrewsi. BAS-AH is a single chain protein and the apparent molecular weight is about 63 kDa as judged by SDS-PAGE. BAS-AH was determined to bind heme (0.89 mol heme/mol protein) as determined by pyridine haemochrome analysis. Fifty percentage cytotoxic concentration (CC50) of BAS-AH on C8166 cells was 9.5 mu M. However, at concentrations that showed little effect oil cell viability, BAS-AH displayed dose dependent inhibition oil HIV-1 infection and replication. The antiviral selectivity indexes corresponding to the measurements of syncytium formation and HIV-1 p24 (CC50/EC50) were 14.4 and 11.4, respectively, corresponding to the . BAS-AH also showed an inhibitory effect on the activity of recombinant HIV-1 reverse transcriptase (IC50 = 1.32 mu M). The N-terminal sequence of BAS-AH was determined to be NAKXKADVIGKISILLGQDNLSNIVAM, which exhibited little identity with other known anti-HIV-1 proteins. BAS-AH is devoid of antibacterial, protcolytic, trypsin inhibitory activity, (L)-amino acid oxidase activity and catalase activity. (c) 2005 Elsevier Ltd. All rights reserved.

Purification and characterization of an irreversible serine protease inhibitor from skin secretions of Bufo andrewsi

Amphibian skin secretions contain many bioactive compounds. In the present work, an irreversible serine protease inhibitor, termed baserpin, was purified for the first time from the skin secretions of toad Bufo andrewsi by Successive ion-exchange and gelfiltration chromatography. Baserpin is a single chain glycoprotein, with an apparent molecular weight of about 60 kDa in SDS-PAGE. Baserpin is an irreversible inhibitor and effectively inhibits the catalytic activity of trypsin, chymotrypsin and elastase. SDS-stable baserpin-trypsin complex could be seen in SDS-PAGE indicates that it possibly belongs to the serpin superfamily. According to the association rates determined, baserpin is a potent inhibitor of bovine trypsin (4.6 X 10(6) M-1 S-1), bovine chymotrypsin (8.9 X 10(6) M-1 s(-1)) and porcine elastase (6.8 X 10(6) M-1 s(-1)), whereas it shows no inhibitory effect on thrombin. The N-terminal sequence of baserpin is HTQYPDILIAKPXDK, which shows no similarity with other known serine protease inhibitors. (c) 2005 Elsevier Ltd. All rights reserved.

Frog albumin is expressed in skin and characterized as a novel potent trypsin inhibitor

A novel potent trypsin inhibitor was purified and characterized from frog Bombina maxima skin. A full-length cDNA encoding the protein was obtained from a cDNA library constructed from the skin. Sequence analysis established that the protein actually comprises three conserved albumin domains. B. maxima serum albumin was subsequently purified, and its coding cDNA was further obtained by PCR-based cloning from the frog liver. Only two amino acid variations were found in the albumin sequences from the skin and the serum. However, the skin protein is distinct from the serum protein by binding of a haem b (0.95 mol/mol protein). Different from bovine serum albumin, B. maxima albumin potently inhibited trypsin. It bound tightly with trypsin in a 1: 1 molar ratio. The equilibrium dissociation constants (K-D) obtained for the skin and the serum proteins were 1.92 x 10(-9) M and 1.55 x 10(-9) M, respectively. B. maxima albumin formed a noncovalent complex with trypsin through an exposed loop formed by a disulfide bond (Cys(53)-Cys(62)), which comprises the scissile bond Arg(58)(P-1)-His(59)(P-1'). No inhibitory effects on thrombin, chymotrypsin, elastase, and subtilisin were observed under the assay conditions. Immunohistochemical study showed that B. maxima albumin is widely distributed around the membranes of epithelial layer cells and within the stratum spongiosum of dermis in the skin, suggesting that it plays important roles in skin physiological functions, such as water economy, metabolite exchange, and osmoregulation.

Anti-infection peptidomics of amphibian skin

Peptidomics and genomics analyses were used to study an anti-infection array of peptides of amphibian skin. 372 cDNA sequences of antimicrobial peptides were characterized from a single individual skin of the frog Odorrana grahami that encode 107 novel an

Isolation and preliminary characterization of a 22-kDa protein with trypsin inhibitory activity from toad Bufo andrewsi skin

A novel trypsin inhibitor termed BATI was purified to homogeneity from the skin extracts of toad Bufo andrewsi by successive ion-exchange, gel-filtration and reverse-phase chromatography. BATI is basic single chain glycoprotein, with apparent molecular weight of 22 kDa in SDS-PAGE. BATI is a thermal stable competitive inhibitor and effectively inhibits trypsin's catalytic activity on peptide substrate with the inhibitor constant (K-i) value of 14 nM and shows no inhibitory effect on chymotrypsin, thrombin and elastase. The N-terminal sequence of BATI is EKDSITD, which shows no similarity with other known trypsin inhibitors. (c) 2005 Elsevier Ltd. All rights reserved.

Multiple bombesin-like peptides with opposite functions from skin of Odorrana grahami

Bombesin-like peptides (BLPs) are a family of neuroendocrinic peptides that mediate a variety of biological activities. Three mature BLPs from the skin secretions of the frog Odorrana grahami were purified. Several bombesin-like peptide cDNA sequences enc

Identification and molecular cloning of a novel neuromedin U analog from the skin secretions of toad Bombina maxima

Amphibian skin contains rich neuropeptides. In the present study, a novel neuromedin U (NmU) analog was isolated from skin secretions of Chinese red belly Load Bombina maxima. Being 17-amino acids long, its primary structure was established as DSSGIVGRPFFLFRPRN-NH2, in which the C-terminal 8-residue segment (FFLFRPRN) is the same as that of rat NmU, while the N-terminal part DSSGIVGRP shows a great sequence variation compared with those of NmU peptides from different resources. The peptide, named Bm-NmU-17, was found to elicit concentration-dependent contractile effects on smooth muscle of rat uterus horns. The cDNA Structure of the peptide, as obtained by a 3'-RACE strategy and subsequently cloning from a skin cDNA library, was found to contain a coding region of 438 nucleotides. The encoded precursor is composed of 145 amino acids with a single copy of Bm-NmU-17 located towards the C-terminus. The sequence of the peptide is preceded by a dibasic site (Lys-Arg) and followed by the sequence of Gly-Arg-Lys, providing the sites of cleavage and releasing of the mature peptide. (c) 2005 Elsevier B.V. All rights reserved.

Bm-TFF2, a trefoil factor protein with platelet activation activity from frog Bombina maxima skin secretions

In mammals, trefoil factor family (TFF) proteins are involved in mucosal maintenance and repair, and they are also implicated in tumor suppression and cancer progression. A novel two domain TFF protein from frog Bombina maxima skin secretions (Bm-TFF2) has been purified and cloned. It activated human platelets in a dose-dependent manner and activation of integrin a(11b)beta(3) was involved. Aspirin and apyrase did not largely reduce platelet response to Bm-TFF2 (a 30% inhibition), indicating that the aggregation is not substantially dependent on ADP and thromboxane A2 autocrine feedback. Elimination of external Ca2+ with EGTA did not influence the platelet aggregation induced by Bm-TFF2, meanwhile a strong calcium signal (cytoplasmic Ca2+ release) was detected, suggesting that activation of phospholipase C (PLC) is involved. Subsequent immunoblotting revealed that, unlike in platelets activated by stejnulxin (a glycoprotein VI agonist), PLC gamma 2 was not phosphorylated in platelets activated by Bm-TFF2. FITC-labeled Bm-TFF2 bound to platelet membranes. Bm-TFF2 is the first TFF protein reported to possess human platelet activation activity. (c) 2005 Elsevier Inc. All rights reserved.