| Sequence analysis and expression of groE gene encoding heat shock proteins of Brucella abortus isolates |
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Tae-Yong Kim1, Ji-Young Kim2, Kyung-Soo Chang3, Myung-Cheol Kim2, Chang-Sik Park4, Hong-Ryul Han5, Moo-Hyung Jun2 |
1ChoongAng Vaccine Laboratory
2College of Veterinary Medicine, Chungnam National University
3College of Medicine, University of Kentucky
4Research Center for Transgenic Cloned Pigs, Chungnam National University
5College of Veterinary Medicine, Seoul National University |
| Brucella abortus 국내 분리주의 Heat Shock Protein 암호 groE 유전자의 염기서열 분석과 발현 |
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김태용1, 김지영2, 장경수3, 김명철2, 박창식4, 한홍율5, 전무형2 |
1(주)중앙백신연구소
2충남대학교 수의과대학
3켄터키대학교 의과대학
4충남대학교 형질전환복제돼지연구센터
5서울대학교 수의과대학 |
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| Abstract |
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GroE that is a heat shock protein composed of GroEL and GroES is known as an immunodominant target of both the humoral and cellular immune responses in bovine brucellosis. This study was carried out to characterize groE gene encoding heat shock proteins of B. abortus isolated in Korea and to evaluate the immunogenicity of the GroE protein expressed in E. coli system. In PCR the specific signals with the size of 2,077 bp were detected in five strains isolated from the mammary lymphnodes of the dairy cattle that were serologically positive and the reference strains. In comparison of the sequences of nucleotides and amino acids among the strains, GroES showed 100% identity in both sequences. GroEL was evaluated 99.0~99.9% in nucleotides and 98.0~100% homology in amino acids. The groE gene including groES and groEL was inserted into pET29a vector and constructed pET29a-GroE recombinant plasmids. The inserted groE was confirmed by digestion with Nco1 and EcoR1 endonucleases and nucleotide sequencing. E. coli BL (DE3) was transformed with pET29a-GroE, named as E. coli BL (DE3)/pET29a-GroE. In SDS-PAGE, it was evident that the recombinant plasmid effectively expressed the polypeptides for GroES (10 kDa) and GroEL (60 kDa) in 0.5, 1 and 2 hours after IPTG induction. The immuno-reactivity of the expressed proteins were proved in mouse inoculation and Western blot analysis. |
| Key Words:
B. abortus, groE gene, nucleotides sequences, expression of groE |
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