scholarly journals Hypothetical Protein Cpn0308 Is Localized in the Chlamydia pneumoniae Inclusion Membrane

2006 ◽  
Vol 75 (1) ◽  
pp. 497-503 ◽  
Author(s):  
Jianhua Luo ◽  
Tianjun Jia ◽  
Rhonda Flores ◽  
Ding Chen ◽  
Guangming Zhong
Keyword(s):  
Membrane Protein ◽  
Fusion Proteins ◽  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Open Reading Frame ◽  
Inclusion Membrane ◽  
Reading Frame ◽  
Antibody Staining ◽  
Infected Cells ◽  
Antibody Labeling

ABSTRACT The hypothetical protein encoded by Chlamydia pneumoniae open reading frame cpn0308 was detected in inclusion membranes of C. pneumoniae-infected cells using antibodies raised with Cpn0308 fusion proteins. The anti-Cpn0308 antibodies did not cross-react with IncA, a known C. pneumoniae inclusion membrane protein, although the anti-Cpn0308 antibody staining overlapped with the anti-IncA antibody labeling. The labeling of the inclusion membrane by the anti-Cpn0308 antibody was specifically blocked by the Cpn0308 but not IncA fusion proteins. The Cpn0308 antigen was detectable 24 h after infection and remained in the inclusion membrane throughout the infection course.

scholarly journals Characterization of the hypothetical protein Cpn1027, a newly identified inclusion membrane protein unique to Chlamydia pneumoniae

Microbiology ◽  
2007 ◽  
Vol 153 (3) ◽  
pp. 777-786 ◽  
Author(s):  
R. Flores ◽  
J. Luo ◽  
D. Chen ◽  
G. Sturgeon ◽  
P. Shivshankar ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Inclusion Membrane

scholarly journals Localization of the Hypothetical Protein Cpn0797 in the Cytoplasm of Chlamydia pneumoniae-Infected Host Cells

2006 ◽  
Vol 74 (11) ◽  
pp. 6479-6486 ◽  
Author(s):  
Feng Dong ◽  
Rhonda Flores ◽  
Ding Chen ◽  
Jianhua Luo ◽  
Youmin Zhong ◽  
...  
Keyword(s):  
Host Cell ◽  
Fusion Proteins ◽  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Host Cells ◽  
Infected Host ◽  
Reading Frame ◽  
The Third ◽  
Cell Cytosol ◽  
Host Cell Cytosol

ABSTRACT Using antibodies raised with chlamydial fusion proteins, we have localized a protein encoded by the hypothetical open reading frame Cpn0797 in the cytoplasm of Chlamydia pneumoniae-infected host cells. The anti-Cpn0797 antibodies specifically recognized Cpn0797 protein without cross-reacting with either CPAFcp or Cpn0796, the only two proteins known to be secreted into the host cell cytosol by C. pneumoniae organisms. Thus, Cpn0797 represents the third C. pneumoniae protein secreted into the host cell cytosol experimentally identified so far.

scholarly journals Localization of the hypothetical protein Cpn0585 in the inclusion membrane of Chlamydia pneumoniae-infected cells

2007 ◽  
Vol 42 (2-3) ◽  
pp. 111-116 ◽  
Author(s):  
Jianhua Luo ◽  
Tianjun Jia ◽  
Youmin Zhong ◽  
Ding Chen ◽  
Rhonda Flores ◽  
...  
Keyword(s):  
Chlamydia Pneumoniae ◽  
Hypothetical Protein ◽  
Inclusion Membrane ◽  
Infected Cells

scholarly journals Identification by Subtractive Hybridization of a Novel Insertion Sequence Specific for Virulent Strains ofPorphyromonas gingivalis

1999 ◽  
Vol 67 (11) ◽  
pp. 5621-5625 ◽  
Author(s):  
Koichi Sawada ◽  
Susumu Kokeguchi ◽  
Hiroshi Hongyo ◽  
Satoko Sawada ◽  
Manabu Miyamoto ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Subtractive Hybridization ◽  
Open Reading Frame ◽  
Specific Marker ◽  
Avirulent Strain ◽  
Target Sequence ◽  
Dna Fragments ◽  
Reading Frame

ABSTRACT Subtractive hybridization was employed to isolate specific genes from virulent Porphyromonas gingivalis strains that are possibly related to abscess formation. The genomic DNA from the virulent strain P. gingivalis W83 was subtracted with DNA from the avirulent strain ATCC 33277. Three clones unique to strain W83 were isolated and sequenced. The cloned DNA fragments were 885, 369, and 132 bp and had slight homology with only Bacillus stearothermophilus IS5377, which is a putative transposase. The regions flanking the cloned DNA fragments were isolated and sequenced, and the gene structure around the clones was revealed. These three clones were located side-by-side in a gene reported as an outer membrane protein. The three clones interrupt the open reading frame of the outer membrane protein gene. This inserted DNA, consisting of three isolated clones, was designated IS1598, which was 1,396 bp (i.e., a 1,158-bp open reading frame) in length and was flanked by 16-bp terminal inverted repeats and a 9-bp duplicated target sequence. IS1598 was detected inP. gingivalis W83, W50, and FDC 381 by Southern hybridization. All three P. gingivalis strains have been shown to possess abscess-forming ability in animal models. However, IS1598 was not detected in avirulent strains of P. gingivalis, including ATCC 33277. The IS1598 may interrupt the synthesis of the outer membrane protein, resulting in changes in the structure of the bacterial outer membrane. The IS1598 isolated in this study is a novel insertion element which might be a specific marker for virulent P. gingivalisstrains.

scholarly journals Nipah Virus Edits Its P Gene at High Frequency To Express the V and W Proteins

2009 ◽  
Vol 83 (8) ◽  
pp. 3982-3987 ◽  
Author(s):  
Sachin Kulkarni ◽  
Valentina Volchkova ◽  
Christopher F. Basler ◽  
Peter Palese ◽  
Viktor E. Volchkov ◽  
...  
Keyword(s):  
High Frequency ◽  
Nipah Virus ◽  
Open Reading Frame ◽  
Mrna Editing ◽  
Reading Frame ◽  
Specific Antibodies ◽  
P Gene ◽  
Gene Transcripts ◽  
Infected Cells ◽  
Over Time

ABSTRACT Nipah virus (NiV) is predicted to encode four proteins from its P gene (P, V, W, and C) via mRNA editing and an alternate open reading frame. By use of specific antibodies, the expression of the V, W, and C proteins in NiV-infected cells has now been confirmed. Analysis of the P-gene transcripts shows a ratio of P:V:W mRNA of 1:1:1, but this differs over time, with greater proportions of V and W transcripts observed as the infection progresses. Eighty-two percent of transcripts are edited, with up to 11 G insertions observed. This exceptionally high editing frequency ensures expression of the V and W proteins.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Open Reading Frame 57 Encodes a Posttranscriptional Regulator with Multiple Distinct Activities

2000 ◽  
Vol 74 (8) ◽  
pp. 3586-3597 ◽  
Author(s):  
Jessica R. Kirshner ◽  
David M. Lukac ◽  
Jean Chang ◽  
Don Ganem
Keyword(s):  
Posttranscriptional Regulation ◽  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Open Reading Frame ◽  
Luciferase Reporter ◽  
Reading Frame ◽  
Infected Cells ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus ◽  
Coding Potential

ABSTRACT Open reading frame (ORF) 57 of Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a homolog of known posttranscriptional regulators that are essential for replication in other herpesviruses. Here, we examined the expression of this gene and the function(s) of its product. KSHV ORF 57 is expressed very early in infection from a 1.6-kb spliced RNA bearing several in-frame initiation codons. Its product is a nuclear protein that, in transient assays, has little effect on the expression of luciferase reporter genes driven by a variety of KSHV and heterologous promoters. However, ORF 57 protein enhances the accumulation of several viral transcripts, in a manner suggesting posttranscriptional regulation. These transcripts include not only known cytoplasmic mRNAs (e.g., ORF 59) but also a nuclear RNA (nut-1) that lacks coding potential. Finally, ORF 57 protein can also modulate the effects of the ORF 50 gene product, a classical transactivator known to be required for lytic induction. The expression from some (e.g., nut-1) but not all (e.g., tk) ORF 50-responsive promoters can be synergistically enhanced by coexpression of ORF 50 and ORF 57. This effect is not due to upregulation of ORF 50 expression but rather to a posttranslational enhancement of the transcriptional activity of ORF 50. These data indicate that ORF 57 is a powerful pleiotropic effector that can act on several posttranscriptional levels to modulate the expression of viral genes in infected cells.

scholarly journals Mouse Hepatitis Virus 3C-Like Protease Cleaves a 22-Kilodalton Protein from the Open Reading Frame 1a Polyprotein in Virus-Infected Cells and In Vitro

1998 ◽  
Vol 72 (3) ◽  
pp. 2265-2271 ◽  
Author(s):  
Xiao Tao Lu ◽  
Amy C. Sims ◽  
Mark R. Denison
Keyword(s):  
Hepatitis Virus ◽  
Mouse Hepatitis Virus ◽  
Open Reading Frame ◽  
Cleavage Sites ◽  
Reading Frame ◽  
Amino Terminal ◽  
Infected Cells ◽  
Carboxy Terminal ◽  
A Site

ABSTRACT The 3C-like proteinase (3CLpro) of mouse hepatitis virus (MHV) is predicted to cleave at least 11 sites in the 803-kDa gene 1 polyprotein, resulting in maturation of proteinase, polymerase, and helicase proteins. However, most of these cleavage sites have not been experimentally confirmed and the proteins have not been identified in vitro or in virus-infected cells. We used specific antibodies to identify and characterize a 22-kDa protein (p1a-22) expressed from gene 1 in MHV A59-infected DBT cells. Processing of p1a-22 from the polyprotein began immediately after translation, but some processing continued for several hours. Amino-terminal sequencing of p1a-22 purified from MHV-infected cells showed that it was cleaved at a putative 3CLpro cleavage site, Gln_Ser4014 (where the underscore indicates the site of cleavage), that is located between the 3CLpro domain and the end of open reading frame (ORF) 1a. Subclones of this region of gene 1 were used to express polypeptides in vitro that contained one or more 3CLpro cleavage sites, and cleavage of these substrates by recombinant 3CLpro in vitro confirmed that amino-terminal cleavage of p1a-22 occurred at Gln_Ser4014. We demonstrated that the carboxy-terminal cleavage of the p1a-22 protein occurred at Gln_Asn4208, a sequence that had not been predicted as a site for cleavage by MHV 3CLpro. Our results demonstrate the usefulness of recombinant MHV 3CLpro in identifying and confirming cleavage sites within the gene 1 polyprotein. Based on our results, we predict that at least seven mature proteins are processed from the ORF 1a polyprotein by 3CLpro and suggest that additional noncanonical cleavage sites may be used by 3CLpro during processing of the gene 1 polyprotein.

scholarly journals Proteolytic Processing of the Open Reading Frame 1b-Encoded Part of Arterivirus Replicase Is Mediated by nsp4 Serine Protease and Is Essential for Virus Replication

1999 ◽  
Vol 73 (3) ◽  
pp. 2027-2037 ◽  
Author(s):  
Leonie C. van Dinten ◽  
Sietske Rensen ◽  
Alexander E. Gorbalenya ◽  
Eric J. Snijder
Keyword(s):  
Serine Protease ◽  
Proteolytic Processing ◽  
Equine Arteritis Virus ◽  
Open Reading Frame ◽  
Cleavage Sites ◽  
Nonstructural Proteins ◽  
Site Mutation ◽  
Reading Frame ◽  
Infected Cells ◽  
Unusual Phenotype

ABSTRACT The open reading frame (ORF) 1b-encoded part of the equine arteritis virus (EAV) replicase is expressed by ribosomal frameshifting during genome translation, which results in the production of an ORF1ab fusion protein (345 kDa). Four ORF1b-encoded processing products, nsp9 (p80), nsp10 (p50), nsp11 (p26), and nsp12 (p12), have previously been identified in EAV-infected cells (L. C. van Dinten, A. L. M. Wassenaar, A. E. Gorbalenya, W. J. M. Spaan, and E. J. Snijder, J. Virol. 70:6625–6633, 1996). In the present study, the generation of these four nonstructural proteins was shown to be mediated by the nsp4 serine protease, which is the main viral protease (E. J. Snijder, A. L. M. Wassenaar, L. C. van Dinten, W. J. M. Spaan, and A. E. Gorbalenya, J. Biol. Chem. 271:4864–4871, 1996). Mutagenesis of candidate cleavage sites revealed that Glu-2370/Ser, Gln-2837/Ser, and Glu-3056/Gly are the probable nsp9/10, nsp10/11, and nsp11/12 junctions, respectively. Mutations which abolished ORF1b protein processing were introduced into a recently developed infectious cDNA clone (L. C. van Dinten, J. A. den Boon, A. L. M. Wassenaar, W. J. M. Spaan, and E. J. Snijder, Proc. Natl. Acad. Sci. USA 94:991–997, 1997). An analysis of these mutants showed that the selective blockage of ORF1b processing affected different stages of EAV reproduction. In particular, the mutant with the nsp10/11 cleavage site mutation Gln-2837→Pro displayed an unusual phenotype, since it was still capable of RNA synthesis but was incapable of producing infectious virus.

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