• Journal of Microbiology and Biotechnology
• /
• 제29권3호
• /
• pp.482-488
• /
• 2019

Porcine circovirus type 2 (PCV2) is the causative agent of postweaning multisystemic wasting syndrome (PMWS) in pigs. Replicase (Rep) proteins are considered essential for viral replication. Capsid (Cap) protein is the primary immunogenic protein that induces protective immunity. Little is known about comparison on the immunogenicity of PCV2 Rep and Cap fusion protein and Cap protein. In the present study, recombinant baculoviruses expressing the Rep-Cap fusion protein (Bac-Rep-Cap) and the Cap protein (Bac-Cap) of PCV2 were constructed and confirmed with western blot and indirect fluorescence assay. Immunogenicities of the two recombinant proteins were tested in mice. The titers of antibodies were determined with a PCV2-specific enzyme-linked immunosorbent assay (ELISA) and a serum neutralization assay. The $IFN-{\gamma}$ response of immunized mice was measured by ELISA. The mice immunized with the Bac-Rep-Cap and Bac-Cap successfully produced Cap-specific immunoreaction. The mice immunized with the Bac-Cap developed higher PCV2-specific neutralizing antibody titers than mice injected with the Bac-Rep-Cap. $IFN-{\gamma}$ in the Bac-Rep-Cap group was increased compared to those in the Bac-Cap group. Vaccination of mice with the Bac-Rep-Cap showed significantly decreased protective efficacy compared to the Bac-Cap. Our findings will indubitably not only lead to a better understanding of the immunogenicity of PCV2, but also improved vaccines.

• 약학회지
• /
• 제39권6호
• /
• pp.676-680
• /
• 1995

The nucleotide sequence of Xbal-Mbol fragment of pKH7, a chloramphenicol-resistant($Cm^{r}$) plasmid isolated from multidrug-resistant S. aureus SA2, has been determined. Xbal-Mbol fragment of pKH7 was found to contain two ORFs. One ORF encoded Rap and the other encoded CAT protein. The deduced amino acid sequences of Rep and CAT of pKH7 were compared to those of pUB112 and pC221. Comparisons revealed that there was one amino acid difference in CAT between pKH7 and pUB112. CAT of pKH7 exhibited 98.6% amino acid identity to that of pC221. In case of Rep proteins, a slightly lower homology of 96.4% and 86.7% in amino acid sequences was observed between pKH7 and pUB112 and between pKH7 and pC221, respectively.

• Journal of Microbiology and Biotechnology
• /
• 제13권6호
• /
• pp.1018-1020
• /
• 2003

The complete 1,486 nucleotide sequence of a cryptic plasmid separated from Lactobacillus bifermentans strain A02 isolated from Kimchi has been determined. The plasmid, designated as pA021, encodes a 33,488 Da putative Rep protein. Based on the sequence similarity, the protein shows homology with coding protein of pRS1, a previously reported plasmid of Oenococcus oeni and the replication initiation protein (Rep) of the Staphylococcal pT181 plasmid family.

• BMB Reports
• /
• 제29권1호
• /
• pp.63-67
• /
• 1996

In order to elucidate the molecular mechanism of plasmid incompatibility of broad host-range plasmid R1162, the plasmid-encoded replication protein RepIB was purified and tested for binding to the 20 bp direct repeat (DR) DNA sequence which is reiterated 3 and 1/2 times within the replicative origin of the plasmid. The RepIB protein specifically binds to the DR DNA. Point mutations in the DR which affect expression of plasmid incompatibility also coordinately affect binding. These results indicate that the incompatibility of broad host-range plasmid R1162 is exerted by the DR DNA by titrating the essential replication protein RepIB.

• Journal of Microbiology and Biotechnology
• /
• 제17권11호
• /
• pp.1841-1847
• /
• 2007

We previously identified the origin of replication of p703/5, a small cryptic plasmid from the KBL703 strain of Enterococcus faecalis. The origin of replication contains putative regulatory cis-elements required for replication and a replication initiator (RepA) gene. The replicon of p703/5 is similar in its structural organization to theta-type plasmids, and RepA is homologous to a family of Rep proteins identified in several plasmids from Gram-positive bacteria. Here, we report molecular interactions between RepA and the replication origin of p703/5. DNase I footprinting using recombinant RepA together with electrophoretic mobility shift assays confirmed the binding of RepA to the replication origin of p703/5 via iterons and an inverted repeat. We also demonstrated the formation of RepA dimers and the different binding of RepA to the iteron and the inverted repeat using gel filtration chromatographic analysis, a chemical crosslinking assay, and electrophoretic mobility shift assays in the presence of guanidine hydrochloride. Our results suggest that RepA plays a regulatory role in the replication of the enterococcal plasmid p703/5 via mechanisms similar to those of typical iteroncarrying theta-type plasmids.

• BMB Reports
• /
• 제34권6호
• /
• pp.584-589
• /
• 2001

One of the cryptic plasmids from the oil degrading bacterium Klebsiella sp. KCL-2, the small plasmid pGD2, has been identified and characterized. This plasmid has a size of 3.6 kb with unknown functions. We constructed the recombinant plasmid pMGD2. The nucleotide sequences of the plasmid were determined and two open reading frames were detected. ORF1 encodes a replication initiator protein (RepA), which has a high degree of homology with the protein of ColE2 plasmid. The product encoded by ORF2 showed a high similarity with the transposase protein of IS5. IS5 is 1195 by long and contains an inverted terminal repetition of 16 bp with one mismatch. Stem-loop structures in the 5'untranslated region of the repA suggest that a putative gene, incA, is located in a complementary strand to the leader region of the repA mRNA.

• 한국동물학회:학술대회논문집
• /
• 한국동물학회 1998년도 한국생물과학협회 학술발표대회
• /
• pp.336.2-336
• /
• 1998

No Abstract, See Full Text

• 생명과학회지
• /
• 제34권2호
• /
• pp.86-93
• /
• 2024

플라스미드의 복제는 엄격하게 조절되어야 하기 때문에 일반적으로 rolling circle 복제를 수행하는 플라스미드들은 복제 개시인자인 RepB는 전사 및 번역 수준에서 엄격하게 조절되어 일정한 copy number를 유지한다. 플라스미드 pJB01에는 단일 오페론으로 구성된 세 개의 orfs (copA, repB, repC 또는 repABC)가 포함되어 있다. 아미노산 서열 분석에서 pJB01 CopA는 다른 플라스미드의 복제 수 조절 단백질로서 Cops와 상동성을 보였다. pMV158의 CopG와 비교할 때, CopA는 플라스미드의 일반화된 억제자의 모티브로 알려진 RHH (ribbon-helix-helix)를 형성하는 것으로 추정된다. gel mobility shift assay 결과 정제된 융합 단백질이 repABC 오페론의 operator 영역에 결합하는 것으로 나타났다. 전사 수준에 대한 CopA의 기능적 역할을 조사하기 위해 CopA R16M, K26R 및 E50V와 같은 세 개의 포인트 돌연변이가 CopA의 코딩 프레임에서 구성되었다. CopA R16M, K26R 및 E50V 돌연변이의 repABC mRNA 수준은 CopA wt보다 각각 1.84, 1.78 및 2.86배 증가했다. 또한 세 개의 CopA 유전자의 돌연변이로 인한 복제 수도 CopA wt보다 각각 1.86, 1.68 및 2.89배 증가했다. 이러한 결과는 CopA가 전사 억제자이며 복제 개시자로서 repABC mRNA 및 RepB 단백질 수를 감소시킴으로써 pJB01의 복제 수를 감소시키는 것으로 제의된다.

• 미생물학회지
• /
• 제31권1호
• /
• pp.44-47
• /
• 1993

A replication initiation gene was identified and its nucleotide sequence has been determined from a 3.8 kb, chloramphenicol acethyltransferase conferring R-plasmid pSBK203 of Staphylococcus aures. Location of the replication related region of pSBK 203 was determined by interuption with pUC 119 at XBaI and MspI sites which resulted in inactivation of replication in Bacilius subtilis. Base sequence of this region revealed on open reading frame of 942 base pairs, which encoded a 314 amino acid protein. Base sequence homology with other rep of pT181 family plasmids such as pT181, pC221, pC223, pS194, pU112, and pCW7 was ranged from 78% to 97% and the predicted amino acid sequence homology was from 72% to 95%.

• 한국결정학회:학술대회논문집
• /
• 한국결정학회 2001년도 임시총회 및 추계학술대회연구발표회(한국결정학회)
• /
• pp.8-8
• /
• 2001