Sequence, structure, and expression of a wasp venom protein with a negatively charged signal peptide and a novel repeating internal structure.
An expression cDNA library prepared from mRNA from the venom gland of a parasitic wasp, Chelonus sp. near curvimaculatus, was screened with polyclonal antibodies against a 33-kDa venom protein from this wasp. Immunoreactive clones were sequenced, yielding a complete inferred sequence for a protein with an NH2 terminus identical with that of the 33-kDa protein. The structure of the cDNA showed an apparent encoded signal peptide, which was unusual in possessing 2 glutamic acid residues juxtapositioned next to, or replacing, the conventional basic residues. The bulk of the mature protein sequence which follows the NH2-terminal, 5000-kDa hydrophobic domain is composed of a dozen tandem repeats of a highly charged, approximately 14-residue sequence, except for the truncated eighth repeat which terminates in the only proline in this large domain. The primary structure is not closely related to any sequence in the GenBank data bank. Secondary structure analysis identified a turn occurring at or near each of 12 invariantly conserved serine residues. Further, the codon used for this serine was invariant, whereas other serines in the protein (including a nearly invariant serine 2 residues away) used various codons. Results of epitope mapping experiments supported a proposed tertiary structure in which the NH2-terminal 5-kDa forms a hydrophobic core, overlain with the charged repeats. Northern analysis of poly(A) RNA from the venom gland of young adult female wasps showed expression of a single 1-kilobase transcript, for which there is no corresponding message in normal or parasitized host larvae. The remarkable structure of this protein and structural data on other wasp venom proteins suggest an evolutionary pattern in which some proteins critical for venom function evolve by internal tandem duplication, and which are secreted after biosynthesis by a different mechanism from that used for proteins with classical signal peptides containing basic residues.
Jones
D
D
Graduate Center for Toxicology, University of Kentucky, Lexington 40506.
Sawicki
G
G
Wozniak
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D004247:DNA; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D014176:Protein Biosynthesis; D021382:Protein Sorting Signals / Q000235:genetics*; D011506:Proteins / Q000235:genetics*; D012313:RNA / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid; D014158:Transcription, Genetic; D014862:Wasp Venoms / Q000378:metabolism*
D021382:Protein Sorting Signals; D011506:Proteins; D014862:Wasp Venoms; D012313:RNA; D004247:DNA