Cloning and expression in Escherichia coli of a Fasciola hepatica gene encoding a calcium-binding protein

被引：25

作者：

de Eguino, ADR

Machín, A

Casais, R

Castro, AM

Boga, JA

Martín-Alonso, JM

Parra, F ^{[1
]}

机构：

[1] Univ Oviedo, Dept Bioquim & Biol Mol, Inst Univ Biotechnol Asturias, CSIC, E-33006 Oviedo, Spain

[2] Inst Nacl Saude, Parasitol Lab, P-4000 Oporto, Portugal

来源：

MOLECULAR AND BIOCHEMICAL PARASITOLOGY | 1999年 / 101卷 / 1-2期

关键词：

Fasciola hepatica; calcium-binding protein; EF-hand; excretory-secretory antigen; helix-loop-helix motif;

D O I：

10.1016/S0166-6851(99)00012-2

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

A Fasciola hepatica cDNA clone of 994 bp was isolated from an adult worm cDNA expression library using a rabbit serum against the excretory-secretory antigens. The nucleotide sequence of the cDNA clone revealed the presence of an open reading frame of 572 bp which encoded a 22 kDa polypeptide (Fh22) showing putative EF-hand domains. This gene was expressed in Escherichia coli and the recombinant protein used for the production of specific antibodies. Immunoblotting studies using the anti-Fh22 serum showed the presence of a polypeptide of similar molecular mass in the excretory-secretory extract of the adult parasite. The recombinant Fh22 polypeptide showed calcium-dependent electrophoretic mobility (decreased with Ca2+-ions and increased with EGTA). The observed behaviour of recombinant Fh22 in gel filtration experiments also suggested calcium-induced conformational changes. (C) 1999 Published by Elsevier Science B.V. All rights reserved.

引用

页码：13 / 21

页数：9

共 34 条

[1] The PROSITE database, its status in 1997 [J].

Bairoch, A ;

Bucher, P ;

Hofmann, K .

NUCLEIC ACIDS RESEARCH, 1997, 25 (01) :217-221

[2] Proteinases secreted by Fasciola hepatica degrade extracellular matrix and basement membrane components [J].

Berasain, P ;

Goni, F ;

McGonigle, S ;

Dowd, A ;

Dalton, JP ;

Frangione, B ;

Carmona, C .

JOURNAL OF PARASITOLOGY, 1997, 83 (01) :1-5

[3]

BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3

[4]

CAWDERY MJH, 1977, BRIT VET J, V133, P145

[5]

COX JA, 1986, J BIOL CHEM, V261, P3173

[6] Characterization of the alpha-ketoglutarate dehydrogenase complex from Fasciola hepatica: Potential implications for the role of calcium in the regulation of helminth mitochondrial metabolism. [J].

Diaz, F ;

Komuniecki, R .

MOLECULAR AND BIOCHEMICAL PARASITOLOGY, 1996, 81 (02) :243-246

[7] CALCIUM-INDUCED STRUCTURAL-CHANGES AND DOMAIN AUTONOMY IN CALMODULIN [J].

FINN, BE ;

EVENAS, J ;

DRAKENBERG, T ;

WALTHO, JP ;

THULIN, E ;

FORSEN, S .

NATURE STRUCTURAL BIOLOGY, 1995, 2 (09) :777-783

[8] CLONING OF A 21.7-KDA VACCINE-DOMINANT ANTIGEN GENE OF SCHISTOSOMA-MANSONI REVEALS AN EF HAND-LIKE MOTIF [J].

FRANCIS, P ;

BICKLE, Q .

MOLECULAR AND BIOCHEMICAL PARASITOLOGY, 1992, 50 (02) :215-224

[9] ECONOMIC AND PRODUCTION CONSEQUENCES OF LIVER FLUKE DISEASE IN DOUBLE-MUSCLED FATTENING CATTLE [J].

GENICOT, B ;

MOULIGNEAU, F ;

LEKEUX, P .

JOURNAL OF VETERINARY MEDICINE SERIES B-ZENTRALBLATT FUR VETERINARMEDIZIN REIHE B-INFECTIOUS DISEASES AND VETERINARY PUBLIC HEALTH, 1991, 38 (03) :203-208

[10]

HAFALLA JCR, IN PRESS MOL BIOCH P

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