title: Cloning and characterization of a cDNA coding for Astacus embryonic astacin, a member of the astacin family of metalloproteases from the crayfish Astacus astacus.
abstract: The astacin family of zinc endopeptidases was named after the digestive enzyme astacin isolated from the crayfish Astacus astacus. Employing a reverse transcription/PCR strategy with degenerate oligonucleotide primers specific for two signature seqences of the astacin family, we have isolated a 1602-bp cDNA from embryos of developing A. astacus eggs, which was designated Astacus embryonic astacin (AEA). This cDNA was found to code for an astacin-like protease domain which accounts for the N-terminal half of the predicted protein. The C-terminal half mainly consists of two complement subcomponent C1r/C1s/embryonic sea urchin protein Uegf/bone morphogenetic protein 1 (CUB) domains. The metalloprotease domain displays an amino acid sequence identity of 42% with astacin. A higher sequence similarity was found to astacin family members that act as hatching enzymes in different species, e.g. chorioallantoic membrane protein 1 (CAM-1; from quail) and Xenopus hatching enzyme (formerly UVS.2), both of which show 54% identity, and high and low choriolytic enzymes (HCE and LCE) from the teleost Oryzias latipes (52% and 48% identity, respectively). A relationship to astacin-like hatching enzymes is further supported by a phylogenetic analysis of the protease domains. Expression of AEA mRNA in developing embryos was found to be restricted to unhatched juveniles (larvae) during the last 8 days before hatching. AEA transcripts could not be detected in various tissues of adult animals or in eggs and embryos from an earlier developmental stage. AEA expression starts about 8 days prior to hatching, followed by a strong (18-fold) induction with a maximum at day 4 before hatching. Newly hatched juveniles were found not to express the AEA mRNA.
authors.affiliation: Zoologisches Institut der Universität Heidelberg, Germany.
authors.forename: G
authors.initials: G
authors.lastname: Geier
authors.forename: R
authors.initials: R
authors.lastname: Zwilling
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003400:Astacoidea / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008666:Metalloendopeptidases / Q000096:biosynthesis* / Q000737:chemistry*; D008969:Molecular Sequence Data; D009990:Oryzias; D016133:Polymerase Chain Reaction; D011784:Quail; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014981:Xenopus
chemical_list: D018076:DNA, Complementary; D011994:Recombinant Proteins; D008666:Metalloendopeptidases; C071288:astacin