Cloning of the cDNA encoding human brain trypsinogen and characterization of its product.
We designed degenerated oligodeoxyribonucleotide primers derived from amino acid (aa) sequences of the highly conserved active sites of mammalian serine proteases (SPs). These primers were used to selectively amplify, in polymerase chain reactions (PCRs), cDNA fragments coding for a SP. We used poly(A)+RNA from human brain to obtain cDNA fragments and amplified one cDNA encoding a novel SP. The full-length nucleotide (nt) sequence was identified by PCR and screening a genomic library in order to obtain the 5'-region. The deduced as sequence shows a high degree of homology to trypsinogens, except for the first exon. In addition to this brain-specific trypsinogen, there exists a variant of the cDNA in pancreas, differing only in the nt sequence of the first exon. An active form of the trypsin was synthesized in vitro and purified by affinity chromatography using soybean trypsin inhibitor (STI) agarose to demonstrate the trypsin-specific interaction with a naturally occurring inhibitor of trypsins.
Wiegand
U
U
Institut für Genetik, Universität zu Köln, Germany.
Corbach
S
S
Minn
A
A
Kang
J
J
Müller-Hill
B
B
D000595:Amino Acid Sequence; D016564:Amyloid beta-Protein Precursor / Q000378:metabolism; D001483:Base Sequence; D001921:Brain / Q000201:enzymology*; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003062:Codon; D017931:DNA Primers; D018076:DNA, Complementary; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014357:Trypsin / Q000235:genetics / Q000302:isolation & purification; D014362:Trypsinogen / Q000235:genetics* / Q000378:metabolism
D016564:Amyloid beta-Protein Precursor; D003062:Codon; D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D014362:Trypsinogen; D012697:Serine Endopeptidases; D014357:Trypsin