Biomedical subjects

Ren Lai

Publications and source records attributed to Ren Lai.

At least 19 recordsLinked to original sources

PubMed2007

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 encoding precursors of BLPs were identified from the skin cDNA library of O. grahami. This is the maximal diversity of BLPs ever found in animals. Five mature BLPs (B1-B5) based on the amino acid sequences derived from the cDNA cloning were synthesized. In the in vitro myotropic contraction experiment, all synthesized BLPs displayed a stimulating effect toward rat stomach strips, except B4 and B5 which showed the opposite effect, suggesting that certain BLPs may act as antagonists of bombesin receptors while most other BLPs act as agonists. This finding will facilitate the finding of novel bombesin receptors and novel ligands of bombesin receptors. The diversity of amphibian BLPs and their precursors were also analyzed and results suggest that amphibian BLPs and corresponding precursors of various sizes and processing patterns can be used as markers of taxonomic and molecular phylogenetics. The remarkable similarity of preproregions gives rise to very different BLPs and 3'-terminal regions in distantly related frog species, suggesting that the corresponding genes form a multigene family originating from a common ancestor. The diversification of BLP loci could thus be part of an evolutionary strategy developed by amphibian species as a result of shifts to novel ecological niches when environmental factors change rapidly.

Amino Acid Sequence↗

PubMed2006

The mastoparanogen from wasp.

Mastoparans are a family of small peptides identified from the venom of hymenopteroid insects. Although they have been characterized as early as 1979, and so far are recognized as a leading biomolecule in potential drug therapy, their precursors, mastoparanogen, have still not been determined. In this paper, several mastoparans from the venom of the wasp Vespa magnifica (Smith) are reported. The cDNA of mastoparanogen is 236 base pairs in length, and encodes 40 amino acid residues, including a N-terminal acidic fragment and a C-terminal mature basic mastoparan, which contain multiple acidic amino acid residues and a tetradecapeptide with three lysines, INLKAIAALAKKLLG, respectively. The glycine at the tetradecapeptide end is the donator of -NH(4) for the amidation of the leucine at the C-terminal. As far as we know, this is the first report of the precursor of animal mastoparan.

Amino Acid Sequence↗

PubMed2006

Amphibian tachykinin precursor.

The precursor of amphibian tachykinin has not been found although more than 30 tachykinins have been isolated from amphibians since 1964. In this report, two tachykinin-like peptides are identified from the skin secretions of the frog, Odorrana grahami. Their amino acid sequences are DDTEDLANKFIGLM-NH(2) (named tachykinin OG1) and DDASDRAKKFYGLM-NH(2) that is the same with ranamargarin found in Rana margaretae, respectively, with a conserved FXGLM-NH(2) C-terminal consensus motif. By cDNA cloning, their precursors were screened from the skin cDNA library of O. grahami. The precursors are composed of 61 amino acid (aa) residues including a signal peptide followed by an acidic spacer peptide and one copy of mature tachykinin-like peptide. Their overall structure is different from structures of other tachykinin precursors such as human protachykinin 1 precursor containing 143 aa including one copy of substance P (SP) and neurokinin A (NKA), and ascidian tachykinin 1 precursor containing 164 aa including two copies of tachykinin-like peptides. The current results demonstrate that the biosynthesis mode of tachykinins in amphibians is different from other animals.

Amino Acid Sequence↗

PubMed2006

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-ALa, b, c, and d, and temporins-ALa, b, and c. The brevinins-AL family which is structurally related to brevinins-1 from skin secretions of the European frog, Rana brevipoda, is composed of 24 amino acids and has an intra-disulfide bridge at the C-terminus. The temporins-AL family, composed of 13 or 16 amino acid residues, is related with temporins from the skin secretions of R. temporaria. The findings of this study will facilitate the solutions to the taxonomic questions of the ranid genus Amolops and Staurois. In the work of this paper, both brevinins-ALb and temporin-Ma induced mast cell degranulation and histamine release, and had cytotoxic activity toward solid tumor cell line HepG(2). Brevinins-ALb also exerted strong hemolytic activity while temporin-Ma had no such activity.

Amino Acid Sequence↗

PubMed2006

The first report of kininogen from invertebrates.

The hornet possesses highly toxic venom, which is rich in toxin, enzymes, and biologically active peptides. Several bradykinin-like peptides, vespakinins, have been found in wasp venoms since 1970s, but the mode of biosynthesis of these peptides is unknown. In the present study, a vespakinin M was purified from venom of Vespa magnifica. Its primary sequence was established as GRPPGFSPFRID. The cDNA encoding the vespakinin M was cloned from the cDNA library of V. magnifica venom gland. The cDNA structure of vespakinin M was found to contain a coding region of 168 nucleotides. The encoded precursor of vespakinin M is composed of a signal peptide, an acidic peptide, and a mature peptide of vespakinin M. This is the first kininogen from insects; it is also the first kininogen from invertebrates. The cDNA structure encoding vespakinin M suggests that the generation mode of bradykinin-related peptides in wasp is different from amphibian skin and mammalian blood system.

Amino Acid Sequence↗

PubMed2006

A novel bradykinin-like peptide from skin secretions of rufous-spotted torrent frog, Amolops loloensis.

A bradykinin-like peptide has been isolated from skin secretions of rufous-spotted torrent frog, Amolops loloensis. This bradykinin-like peptide was named amolopkinin. Its primary structure, RAPVPPGFTPFR, was determined by Edman degradation and mass spectrometry. It is structurally related to bradykinin-like peptides identified from skin secretions of other amphibians. Amolopkinin is composed of 12 amino acid residues and is related to bradykinin composed of nine amino acid residues, identified from the skin secretions of Odorrana schmackeri. Amolopkinin was found to elicit concentration-dependent contractile effects on isolated guinea pig ileum. cDNA clones encoding the precursor of amolopkinin were isolated by screening a skin cDNA library of A. loloensis and then sequenced. The amino acid sequences deduced from the cDNA sequences match well with the results from Edman degradation. Analysis of different amphibian bradykinin cDNA structures revealed that a deficiency of an18-nucleotide fragment (TCAAGAATGATCAGACGC in the cDNA encoding bradykinin from O. schmackeri) in the peptide-coding region resulted in absence of a di-basic site for trypsin-like proteinases and an unusual - APV - insertion in the N-terminal part of amolopkinin. This is the first report of a bradykinin-like peptide comprised of bradykinin with an insertion in its N-terminal part. Our results demonstrate the hypervariability of amphibian bradykinin-like peptides, as well as the diversity of antimicrobial peptides in amphibians.

Amino Acid Sequence↗

PubMed2006

A tick B-cell inhibitory protein from salivary glands of the hard tick, Hyalomma asiaticum asiaticum.

Some studies done to date suggest that B-cell inhibitory factor occurred in tick saliva. In this study, a novel protein having B-cell inhibitory activity was purified and characterized from the salivary glands of the hard tick, Hyalomma asiaticum asiaticum. This protein was named B-cell inhibitory factor (BIF). The cDNA encoding BIF was cloned by cDNA library screening. The predicted protein from the cDNA sequence is composed of 138 amino acids including the mature BIF. No similarity was found by Blast search. The lipopolysaccharide-induced B-cell proliferation was inhibited by BIF. This is the first report of the identification and characterization of B-cell inhibitory protein from tick. The current study facilitates the study of identifying the interaction among tick, Borrelia burgdorferi, the causative agent of Lyme disease, and host.

Amino Acid Sequence↗

PubMed2006

Two antimicrobial peptides from skin secretions of Rana grahami.

Two antimicrobial peptides manifested a broad spectrum of antimicrobial 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 structures are GLLSGILGAGKNIVCGLSGLC and GLLSGILGAGKHIVCGLSGLC, respectively, determined by Edman degradation and mass spectrometry. They are structurally related to nigrocins identified from skin secretions of the dark-spotted frog, Rana nigromaculata. The cDNA clones encoding the precursor of grahamins were screened and sequenced from the skin cDNA library of R. grahami. The amino sequences deduced from the cDNA sequences match well with the results from Edman degradation. As other antimicrobial peptides from Rana species, grahamins contain a C-terminal loop region delineated by an intra-disulfide bridge named Rana box. Based on structural comparison of grahamin with other known antimicrobial peptides, grahamins could be classified into the family of antimicrobial peptides containing a single intra-disulfide bridge.

Animals↗

PubMed2006

Massilia dura sp. nov., Massilia albidiflava sp. nov., Massilia plicata sp. nov. and Massilia lutea sp. nov., isolated from soils in China.

Four Gram-negative, motile, rod-shaped bacterial strains were isolated from soil samples collected from south-east China. A taxonomic study including phylogenetic analysis based on 16S rRNA gene sequences and phenotypic characteristics was performed. DNA G+C contents of the four strains were 63-66 mol%. Their predominant ubiquinone was Q-8. The fatty acid profiles contained C16:1omega7c (36.9-54.7%) and C16:0 (22.8-25.5%) as the major components. Based on their phenotypic characteristics, phylogenetic position as determined by 16S rRNA gene sequence analysis and DNA-DNA hybridization results, the four isolates are considered to represent four novel species of the genus Massilia, for which the names Massilia dura sp. nov. (type strain 16T=CCTCC AB 204070T=KCTC 12342T), Massilia albidiflava sp. nov. (type strain 45T=CCTCC AB 204071T=KCTC 12343T), Massilia plicata sp. nov. (type strain 76T=CCTCC AB 204072T=KCTC 12344T) and Massilia lutea sp. nov. (type strain 101T=CCTCC AB 204073T=KCTC 12345T) are proposed.

Base Composition↗

PubMed2005

[Isolation and purification of an inhibitor on platelet aggregation from Ixodes sinensis].

OBJECTIVE: To study the bioactive components in ticks which inhibit platelet aggregation, and to understand the molecular mechanism of tick-host interaction. METHODS: Sephadex G-50 gel filtration and high performance liquid chromatography (HPLC) were used to purify the platelet aggregation inhibitor from lxodes sinensis. Its molecular weight and purity were checked by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. Platelet-rich plasma (PRP) of rabbit was used to examine the function of platelet aggregation inhibitor. RESULTS: A purified platelet aggregation inhibitor was identified from L. sinensis with a molecular weight of 8 065. It inhibited platelet aggregation induced by ADP with strong potency. The inhibition of platelet aggregation reached over 90% under a concentration of 10 microg/ml. CONCLUSION: An inhibitor of platelet aggregation from L. sinensis was identified, which may play an important role for ticks to successfully get blood meal from their hosts.

Animals↗

PubMed2005

Two families of antimicrobial peptides from wasp (Vespa magnifica) venom.

The hornet possesses highly toxic venom, which is rich in toxin, enzymes and biologically active peptides. Many bioactive substances were identified from wasp venom. Two families of antimicrobial peptides were purified and characterized from the venom of the wasp, Vespamagnifica (Smith). The primary structures of these peptides are homologous to those of chemotactic peptides and mastoparans isolated from other vespid venoms. They also share similarity to temporins which are amphibian antimicrobial peptides identified from the skin of the frog, Ranaboylii. These peptides show antimicrobial activities against bacteria and fungi. However, they show little hemolytic activity against human blood red cells.

Amino Acid Sequence↗

PubMed2005

Direct antimicrobial activities of PR-bombesin.

PR-bombesin is a bombesin-like peptide derived from the skin of the Chinese red belly toad, Bombina maxima. The 8-residue segment of N-terminal of RP-bombesin, comprising four prolines and three basic residues, is extensively different from other bombesin-like peptides. Since sequence of Pro-Arg-Pro generally plays an important role in the antimicrobial activity of proline-rich antimicrobial peptides, the componential feature of PR-bombesin indicates that it may have antimicrobial activity. In this paper, we presented the first evidence that bombesin-like peptides possess direct antimicrobial activities as some neuropeptides. It was determined by CD spectroscopy that PR-bombesin adopted a combination of random coil and beta-sheet structure, suggesting RP-bombesin is a new member of antimicrobial peptides having beta structure but without disulfide bonds. Current results also supported that PR-bombesin plays a direct defensive role besides its neuro-endocrological functions.

Animals↗

PubMed2005

Maximin 9, a novel free thiol containing antimicrobial peptide with antimycoplasma activity from frog Bombina maxima.

Amphibian skin is a rich resource of antimicrobial peptides, like maximins and maximin Hs from frog Bombina maxima. Novel cDNA clones encoding a precursor protein, which comprises a novel maximin peptide (maximin 9) and reported maximin H3, were isolated from two constructed skin cDNA libraries of B. maxima. The predicted primary structure of maximin 9 is GIGRKFLGGVKTTFRCGVKDFASKHLY-NH2. A surprising substitution is at position 16, with a free cysteine in maximin 9 rather than usual conserved glycine in other reported maximins. Maximin 9, the homodimer form and its Cys16 to Gly16 mutant were synthesized and their antimicrobial activities were evaluated. Unlike previously reported maximin 3, the tested bacterial and fungal strains were resistant to maximin 9, its homodimer and the Cys16 to Gly16 mutant (with MICs>100 microM). On the other hand, interestingly, while eight clinical Mollicutes strains were generally resistant to maximin 9 homodimer and its Cys16 to Gly16 mutant, most of them are sensitive to maximin 9 at a peptide concentration of 30 microM, especially in the presence of dithiothreitol. These results indicate that the presence of a reactive Cys residue in maximin 9 is important for its antimycoplasma activity. The diversity of antimicrobial peptide cDNA structures encountered in B. maxima skin cDNA libraries and the antimicrobial specificity differences of the peptides may reflect well the species' adaptation to the unique microbial environments.

Animals↗

PubMed2005

A novel antimicrobial peptide from salivary glands of the hard tick, Ixodes sinensis.

A novel antimicrobial peptide named as ixosin was isolated from the salivary glands of the hard tick, Ixodes sinensis, by gel filtration, ion exchange chromatography and reverse-phase high-performance liquid chromatography (RP-HPLC). Its amino acid sequence was determined as GLHKVMREVLGYERNSYKKFFLR by Edman degradation and its molecular weight was 2870.5 analyzed by fast atom bombardment (FAB) mass spectrometry. This is the first antimicrobial peptide from ticks that lacks cysteine in its primary structure. The cDNA encoding ixosin was cloned by cDNA library screening. The predicted protein from the cDNA sequence composed of 79 amino acids including mature ixosin. Purified ixosin exerted its antimicrobial activities against bacteria and fungi. No similarity was found by BLAST search to any database entries and, thus, our findings describe a novel antimicrobial peptide.

Amino Acid Sequence↗

PubMed2005

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 (KD) 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 (Cys53-Cys62), which comprises the scissile bond Arg58(P1)-His59(P1'). 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.

3' Untranslated Regions↗

PubMed2005

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 toad 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.

Amino Acid Sequence↗

PubMed2005

Variety of antimicrobial peptides in the Bombina maxima toad and evidence of their rapid diversification.

Antimicrobial peptides secreted by the skin of many amphibians play an important role in innate immunity. From two skin cDNA libraries of two individuals of the Chinese red belly toad (Bombina maxima), we identified 56 different antimicrobial peptide cDNA sequences, each of which encodes a precursor peptide that can give rise to two kinds of antimicrobial peptides, maximin and maximin H. Among these cDNA, we found that the mean number of nucleotide substitution per non-synonymous site in both the maximin and maximin H domains significantly exceed the mean number of nucleotide substitution per synonymous site, whereas the same pattern was not observed in other structural regions, such as the signal and propiece peptide regions, suggesting that these antimicrobial peptide genes have been experiencing rapid diversification driven by Darwinian selection. We cloned and sequenced seven genes amplified from skin or liver genomic DNA. These genes have three exons and share the same gene structure, in which both maximin and maximin H are encoded by the third exon. This suggests that alternative splicing and somatic recombination are less likely to play a role in creating the diversity of maximins and maximin Hs. The gene trees based on different domain regions revealed that domain shuffling or gene conversion among these genes might have happened frequently.

Amino Acid Sequence↗

PubMed2004

An opioid peptide from synganglia of the tick, Amblyomma testindinarium.

An opioid peptide, which shares similarity with mammalian hemorphins, has been identified from the synganglia (central nervous system) of the hard tick, Amblyomma testindiarium. Its primary sequence was established as LVVYPWTKM that contains a tetrapeptide sequence Tyr-Pro-Trp-Thr of hemorphin-like opioid peptides. By hot-plate bioassay, the purified peptide and synthetic peptide displayed dose-related antinociceptive effect in mice, as observed for other hemorphin-like opioid peptides. This is the first opioid peptide identified from ticks. Ticks may utilize the opioid peptide in their strategy to escape host immuno-surveillance as well as in inhibiting responses directed against themselves.

Amino Acid Sequence↗