Xian-Chun Zeng

Molecular cloning and sequencing of two 'short chain' and two 'long chain' K(+) channel-blocking peptides from the Chinese scorpion Buthus martensii Karsch.

Five full‐length cDNAs encoding the precursors of two ‘short chain’ scorpion non‐toxic peptides active on Ca2+‐activated K+ channels (BmP02 and BmP03) and two novel putative long chain K+ channel‐blocking peptides (named BmTXKβ and BmTXKβ2) were first isolated from the venom gland cDNA library of the Chinese scorpion Buthus martensii Karsch (BmK). BmTXKβ2 showed a high similarity with AaTXKβ, while BmTXKβ was completely different in the deduced primary structure from the long chain and short chain scorpion toxins already characterized. Thus, BmTXKβ expands the scorpion long chain K+ channel‐blocking peptide family. Although little sequence similarity exists between the above two short and two long peptides, they are similar at the positions of six cysteines, suggesting that they should all share a similar scaffold composed of an α‐helix and a three‐stranded β‐sheet.

Cloning and characterization of a cDNA sequence encoding the precursor of a chlorotoxin-like peptide from the Chinese scorpion Buthus martensii Karsch☆

A full-length cDNA sequence encoding the precursor of a venom peptide with homology to chlorotoxin (named BmKCT) was isolated from a cDNA library made from the venom glands of the Chinese Scorpion Buthus martensii Karsch. The encoded precursor of BmKCT was 59 amino acid residues long including a signal peptide of 24 residues and a mature toxin of 35 residues with four disulfide bridges. The sequence of BmKCT is similar (68% identities) to that of chlorotoxin isolated from Leiurus quinguestriatus quinquestriatus. BmKCT is the first report of the cDNA sequences encoding four-disulfide-bridged short-chain toxins from Buthus martensuii Karsch so far.

Cloning and Characterization of a Novel cDNA Sequence Encoding the Precursor of a Novel Venom Peptide (BmKbpp) Related to a Bradykinin‐Potentiating Peptide from Chinese Scorpion Buthus martensii Karsch

Based on the amino acid sequence of a bradykinin‐potentiating peptide (Bpp) (peptide K‐12) from scorpion Buthus occitanus, a full‐length cDNA sequence encoding the precursor of a novel venom peptide (named BmKbpp) related to this Bpp, has been isolated and analyzed. The cDNA encodes a precursor of 72 amino acid residues, including a signal peptide of 22 residues and an extra Arg‐ArgArg tail at the C‐terminal end of the precursor, which have to be removed in the processing step. The C‐terminal region (21 residues) of the precursor is homologous (57% identical) with the sequence of peptide K‐12. Thus, according to the primary structure of the BmKbpp precursor, there may be a propeptide between the signal peptide and the putative mature BmKbpp at the C‐terminal region of the precursor.

Molecular dissection of venom from Chinese scorpion Mesobuthus martensii: identification and characterization of four novel disulfide-bridged venom peptides.

Scorpion venom is composed of a large repertoire of biologically active polypeptides. However, most of these peptides remain to be identified and characterized. In this paper, we report the identification and characterization of four novel disulfide-bridged venom peptides (named BmKBTx, BmKITx, BmKKx1 and BmKKx2, respectively) from the Chinese scorpion, Mesobuthus martensii (also named Buthus martensii Karsch). BmKBTx is composed of 58 amino acid residues and cross-linked by three disulfide bridges. The sequence of BmKBTx shows some similarities to that of the toxin, birtoxin, and its analogs. It is likely that BmKBTx is a beta-toxin active on Na+ channels, which is toxic to either insects or mammals. BmKITx is composed of 71 amino acid residues with four disulfide bridges. It is the longest venom peptide identified from M. martensii so far. BmKITx shows little sequence identity with scorpion alpha-toxins toxic to insects. It is likely that BmKITx is a new type of Na+ -channel specific toxin active on both insects and mammals. BmKKx1 contains 38 amino acid residues cross-linked by three disulfide bridges and shows 84% sequence identity with BmTx3, an inhibitor of A-type K+ channel and HERG currents. BmKKx1 has been classified as alpha-KTx-15.8. BmKKx2 is composed of 36 residues and stabilized by three disulfide bridges. BmKKx2 is a new member of the gamma-K+ -channel toxin subfamily (classified as gamma-KTx 2.2). The venoms of scorpions thus continue to provide novel toxins with potential novel actions on targets.

Molecular cloning and characterization of four scorpion K+-toxin-like peptides: A new subfamily of venom peptides (α-KTx14) and genomic analysis of a member**

Four full-length cDNAs encoding the precursors of four K(+)-toxin-like peptides (named BmKK(1), BmKK(2), BmKK(3) and BmmKK(4), respectively) were first isolated from a venom gland cDNA library of the Chinese scorpion Buthus martensii Karsch. The deduced precursors of BmKK(1), BmKK(2) and BmKK(3) are all made of 54 amino acid residues including a signal peptide of 23 residues, and a mature toxin of 31 residues with three disulfide bridges. The precursor of BmKK(4) is composed of 55 amino acid residues including a signal peptide of 23 residues, a mature toxin of 30 residues cross-linked by three disulfide bridges, and an extra Gly-Lys tail which should be removed in the processing step. The four peptides displayed 24-97% sequence identity with each other, and less than 27% homology with any other scorpion toxins described. However, they shared a common disulfide bridge pattern, which was consistent with that of most short-chain K(+)-toxins, suggesting they represent a new class of scorpion toxins and their target receptors may be a subfamily of K(+) channels. We classified the BmKK toxin subfamily as alpha-KTx14 according to the classification rules. The genomic sequence of BmKK(2) was also cloned and sequenced. It consisted of two exons, disrupted by an intron of 79 bp inserted in the region encoding the C-terminal part of the signal peptide. This structure was very similar to that of other K(+)-toxins described previously.

Nine novel precursors of Buthus martensii scorpion α-toxin homologues.

Abstract The cDNAs encoding nine novel α-toxin homologues were isolated from the venom gland cDNA library of the Chinese scorpion Buthus martensii Karsch (BmK). They are rich in AAAA and TTTT elements at the 5′ UTRs. The flanking region of the translation initiation codon ATG is AAAATGAA, which is highly conserved in scorpion Na+, K+ and Cl− channel toxin genes. These putative scorpion α-toxins shared 45.5–98.4% homology with the characterized BmK α-toxins, and were completely conserved in the positions of all eight cysteines. This showed, together with higher homology at nucleotide level than that at amino acid level, that these toxins may originate from a common ancestor. The discovery of a series of homologues of scorpion α-toxin with a different degree of natural mutation in the primary structure will provide us with a valuable system for studying the structure–function relationship of scorpion toxins.

Evidence for the existence of insect defensin-like peptide in scorpion venom.

Insect defensin refers to a group of antibacterial peptides derived from a variety of insect species as well as from scorpion and possessing a three‐dimensional structure highly similar to that of scorpion toxins. A full‐length cDNA encoding an insect defensin‐like peptide was isolated from the venom gland cDNA library of the Chinese scorpion

Precursor of a Novel Scorpion Venom Peptide (BmKn1) with no Disulfide Bridge from Buthus martensii Karsch

A full‐length cDNA sequence encoding the precursor of a novel venom peptide (named BmKn1) with no disulfide bridge was first isolated from the venom gland cDNA library of Chinese scorpion Buthus martensii Karsch. The encoded precursor consisted of 70 amino acid residues including two parts: a signal peptide of 23 residues, and a putative mature venom peptide (BmKn1) of 47 residues. The sequence of BmKn1 showed no similarity to those of other scorpion venom peptides. BmKn1 may be the first member of a new venom peptide family from scorpion. Future research will be interesting to unravel further the pharmacological function of this novel scorpion venom peptide.

Molecular characterization of a K+ channel blocker in the scorpionButhus martensii Karsch

K+ channel blockers of scorpion venoms are of important value in studying pharmacology and physiology of specific K+ channel of cells. Based on the amino acid sequences of BmP01 previously characterized as a small-conductance Ca2+-activated K+ channel blocker, two “back to back” degenarate primers have been designed and synthesized for inverse PCR strategy, its full-length cDNA has been cloned from the venom gland of the Chinese scorpionButhus martensii. The cDNA is composed of 3 parts: 5′ UTR, ORF and 3′ UTR. The flanking sequence of translation initiation codon ATG is AAAATGA, which is highly conserved in scorpion Na+ channel toxin and protozoan genes, suggesting that these genes may have followed a common mechanism for translation initiation. The 3′ UTR contains poly(A) signal AATAAA. The open reading frame encodes a precursor of 57 residues with a signal peptide of 28 residues and a mature peptide of 29 residues. The signal peptide is rich in hydrophobic amino acid residues and its length is significantly different from that of the determined scorpion Na+ channel toxin. The deduced amino acid sequence of mature peptide is completely consistent with BmP01 previously determined by primary structure analysis.