Purification and characterization of fibroblast-activating factor isolated from Porphyromonas gingivalis W50

Mihara, J; Holt, Stanley C.

doi:10.1128/iai.61.2.588-595.1993

Cited by 21 publications

(12 citation statements)

References 45 publications

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“…These improvements made it possible to isolate substantial amounts (>30 mg per liter of the culture) of highly purified protein and facilitated a more detailed protein analysis. In this study, we characterized two HmuY variants: a mature HmuY protein (mHmuY) lacking 21 N-terminal amino acids and a mature processed HmuY (mpHmuY) lacking 25 N-terminal amino acids, which were not found in a native protein purified from P. gingivalis cells (Mihara and Holt 1993a;Lewis et al 2006). Since both HmuY variants showed identical properties, except for the slower migration of mpHmuY during electrophoresis under native conditions resulting from the different amino-acid composition, in this study we showed results for mpHmuY only.…”

Section: Biochemical Analysis Of Hmuy Proteinmentioning

confidence: 99%

Porphyromonas gingivalis HmuY and HmuR: further characterization of a novel mechanism of heme utilization

et al. 2007

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Porphyromonas gingivalis HmuY is a putative heme-binding lipoprotein associated with the outer membrane. It is part of an operon together with a gene encoding an outer-membrane hemin utilization receptor (HmuR) and four uncharacterized genes. A similar operon organization was found in Bacteroides fragilis and B. thetaiotaomicron, with the former containing an additional HmuY homologue encoded upstream of the hmuR-like gene. In P. gingivalis cultured under heme-limited conditions, a approximately 1-kb hmuY transcript was produced at high levels along with some approximately 3.5 and approximately 9-kb transcripts. Compared with the parental strain, mutants deficient in hmuY or hmuR or hmuY-hmuR gene function grew more slowly and bound lower amounts of hemin and hemoglobin. Significantly, they grew more slowly or were unable to grow when human serum was used as the sole iron/heme source. Analysis of the hmu promoter showed that it is regulated by iron. The HmuY protein normally occurs as a homodimer, but in the presence of hemin it may form tetramers. These results show that HmuY may be the first reported member of a new class of proteins in Porphyromonas and Bacteroides species involved in heme utilization, a function being exerted in conjunction with HmuR, an outer-membrane heme transporter.

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Section: Biochemical Analysis Of Hmuy Proteinmentioning

confidence: 99%

Porphyromonas gingivalis HmuY and HmuR: further characterization of a novel mechanism of heme utilization

et al. 2007

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“…Progress in molecular biological techniques has resulted in our increased interest in proteins which are directly encoded by DNA. Recently, a number of novel proteins specific to P. gingivalis have been identified and thoroughly investigated, i.e., proteases (2,7,11,31), a fibroblast-activating factor (19), heat shock protein (18), etc. However, the roles of these proteins in vivo remain speculative, and their usefulness as diagnostic tools has not been established.…”

Section: Discussionmentioning

confidence: 99%

A Novel Approach for Detecting an Immunodominant Antigen ofPorphyromonas gingivalisin Diagnosis of Adult Periodontitis

Kawai

Itô

Sakato

et al. 1998

Clin Diagn Lab Immunol

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In the course of long-term infection with Porphyromonas gingivalis in adult periodontitis, a specific antibody response to this organism is generated. We describe a potential novel approach for identifying an immunodominant antigen in human periodontitis patients. First, various monoclonal antibodies (MAbs) were established from mice immunized with crude antigen preparations of P. gingivalis FDC 381. The antigen specificities of these MAbs were compared with those of serum antibodies of 10 periodontitis patients in a competitive enzyme-linked immunosorbent assay. The binding of one MAb (termed PF18) was readily inhibited by sera from all patients but not by sera from healthy volunteers. The antigen recognized by PF18 existed on the cell surface, presumably in the capsule layer, shown by immunoelectron microscopic analysis. Purification of the antigenic substance, termed PF18-Ag, was performed by immunoaffinity chromatography with the MAb. Characterization of PF18-Ag suggested that the epitope was composed of carbohydrates but not peptides and that the substance was different from lipopolysaccharide. Measurement of levels of serum antibody to PF18-Ag better discriminated periodontitis patients from healthy individuals than measurement of antibodies to crude antigen preparations of P. gingivalis. Immunoglobulin G2 was the predominant isotype among the antibodies to PF18-Ag in the patients’ sera. These results suggest that PF18-Ag, which is possibly a novel substance, is an important antigenic substance and is potentially useful for the clinical diagnosis of adult periodontitis. The approach that was used would also be relevant to detecting immunodominant antigens of other infectious microorganisms.

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“…Furthermore, Mihara and Holt (27) isolated a 24-kDa protein from the outer membrane vesicles of P. gingivalis and found that the protein stimulated the proliferation of human gingival fibroblasts. These findings suggest that fibroblasts as well as lymphoid cells proliferate and produce various inflammatory cytokines in response to the cell surface components of black-pigmented anaerobes in periodontal tissues.…”

Section: Discussionmentioning

confidence: 99%

Immunobiological activities of a 55-kilodalton cell surface protein of Prevotella intermedia ATCC 25611

et al. 1994

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A protein was extracted from whole cells of Prevoteila intermedia ATCC 25611 with sodium lauroylsarcosine and purified by chromatography on a DEAE-Sepharose fast-flow column. The apparent molecular weight of the protein was 55,000. A mouse polyclonal antibody specific for the protein recognized the cell surface structure of P. intermedia and also reacted with proteins in lysates of other black-pigmented anaerobic bacteria, such as Porphyromonas endodontalis and Prevotella melaninogenica, but not with those in lysates of Porphyromonas gingivalis or with the purified fimbriae of P. gingivalis 381. The N-terminal sequence of the 55-kDa protein showed only low homology with the cell surface proteins of any black-pigmented bacteria reported to date. The level of immunoglobulin G antibody to the antigen was higher in the sera of patients with periodontitis than in the sera of healthy volunteers. The protein induced interleukin-la,-1pI,-6, and-8 and tumor necrosis factor alpha in human peripheral blood mononuclear cell cultures and interleukin-li and-6 in human umbilical vascular endothelial cell and gingival fibroblast cultures. The protein induced interleukin-6 and tumor necrosis factor alpha activities in peritoneal macrophages from C3H/HeJ as well as from C3H/HeN mice and also induced cytokine activities in the sera of both strains of mice primed with muramyldipeptide.

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Purification and characterization of fibroblast-activating factor isolated from Porphyromonas gingivalis W50

Cited by 21 publications

References 45 publications

Porphyromonas gingivalis HmuY and HmuR: further characterization of a novel mechanism of heme utilization

Porphyromonas gingivalis HmuY and HmuR: further characterization of a novel mechanism of heme utilization

A Novel Approach for Detecting an Immunodominant Antigen ofPorphyromonas gingivalisin Diagnosis of Adult Periodontitis

Immunobiological activities of a 55-kilodalton cell surface protein of Prevotella intermedia ATCC 25611

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