Molecular cloning, sequencing, and expression of cDNA encoding serine protease with fibrinolytic activity from earthworm

Citation
M. Sugimoto et N. Nakajima, Molecular cloning, sequencing, and expression of cDNA encoding serine protease with fibrinolytic activity from earthworm, BIOS BIOT B, 65(7), 2001, pp. 1575-1580
Citations number
29
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Agricultural Chemistry","Biochemistry & Biophysics
Journal title
BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY
ISSN journal
0916-8451 → ACNP
Volume
65
Issue
7
Year of publication
2001
Pages
1575 - 1580
Database
ISI
SICI code
0916-8451(200107)65:7<1575:MCSAEO>2.0.ZU;2-L
Abstract
An earthworm, Lumbricus rubellus, produces alkaline trypsin-like proteases that are greater than trypsins in their stability and strong tolerance to o rganic solvents. cDNAs encoding strong fibrinolytic proteases (F-III-2 and F-III-1) in the six isozymes were cloned and sequenced to study their stabi lity-structure relationship. The cDNAs of F-III-2 and F-III-1 comprised 101 1 and 973 bp and included open reading frames that encode polypeptides of 2 45 and 246 amino acid residues, respectively. The deduced amino acid sequen ces of F-III-2 and F-III-1 have 7 and 8 activation peptides in the N-termin i respectively, indicating that they are translated as proenzymes and modif ied to active forms by posttranslational processing. They showed similarity to mammalian serine proteases and conserved the catalytic amino acid resid ues, however, neither arginine nor lysine residues were present in the auto lysis region. The gene encoding the native form of F-III-2 was expressed in Pichia pastoris to produce and secrete the earthworm protease in the cultu re medium, which dissolves an artificial fibrin plate.