Nucleotide and protein sequences of a proteinase inhibitor from the vitelline envelope of dace (Tribolodon hakonensis) eggs.
Our experimental purpose is to probe the structure(s) of the chorionic proteinase inhibitor and its cDNA sequence(s) and to develop the application of safe medicines for protection of human and other animal bodies from pathogenic microbe attacks. In this study, chorionic proteinase inhibitor protein was isolated, sequenced and used to base the design of PCR primers, which were then used to amplify DNA using RT-PCR. A cDNA clone of the protein which inhibited the activities of serine proteinases and thermolysin was obtained on the basis of mRNA extracted from ovarian tissue of dace, Tribolodon hakonensis, and the deduced amino acid sequence was determined. Chorionic proteinase inhibitor (TribSPI) peptides of about 9.0 kDa (TribSPI) and 14 kDa (TribSPI-S) were purified from vitelline envelope extracts by thermolysin-immobilized affinity-chromatography. The cloned TribSPI cDNA was 1806 bp in length, and the open reading flame (ORF) was 915 bp encoding a protein of 305 amino acid residues. The inhibitor protein had a molecular mass of 33,550 daltons and was composed of five similar domains. Each domain contained eight cysteine residues, and it's deduced amino acid sequence was only 33 approximately 34% identical to those of human and porcine antileukoproteinases (hALP and pALP, respectively). A possible binding-site for serine proteinases, Arg-Ile, was contained in three domains.
Hosomi
Osamu
O
Department of Health Science, School of Health and Sports Science, Juntendo University 1-1, Hiragagakuendai, Inba-mura, Inba-gun, Chiba, Japan. hosomi@sakura.juntendo.ac.jp
Ohe
Yoshihide
Y
Takeya
Akira
A
Hosaka
Kohei
K
Okubo
Toyoji
T
Iyobe
Shizuko
S
Kudo
Shigeharu
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002846:Chromatography, Affinity; D003530:Cyprinidae / Q000378:metabolism*; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D015842:Serine Proteinase Inhibitors / Q000235:genetics* / Q000378:metabolism; D014817:Vitelline Membrane / Q000737:chemistry*
D017931:DNA Primers; D018076:DNA, Complementary; D015842:Serine Proteinase Inhibitors