V antigen-polyhistidine fusion peptide: Binding to LcrH and active immunity against plague

Vladimir Motin, Yuri A. Nedialkov, Robert R. Brubaker

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

The structural gene for V antigen (lcrV) is known to be encoded within the lcrGVH-yopBD operon of the -70-kb low-calcium-response or Lcr plasmid of Yersinia pestis. This 37-kDa monomeric peptide was reported to provide active immunity in mice, suppress inflammatory cytokines, and regulate expression of the low calcium response (Lcr+). Here we describe pVHB62, encoding a polyhistidine-V antigen fusion peptide (V(h)) and linked LcrH. V(h) underwent degradation from both the C terminus and N terminus during classical chromatographic fractionation but remained intact within two compartments during Ni2+ affinity chromatography. The first was homogeneous, capable of active immunization (mouse intravenous 50% lethal dose, > 107 bacteria), and stable at 4°C. The second remained bound to the affinity column but could be eluted as a mixture of V(h), LcrH, and low-molecular-weight material by application of 6 M guanidine · HCl. This mixture was dialyzed, denatured in 8 M urea, and again applied to the affinity column, which then hound V(h) but not LcrH. The latter was recovered and renatured, and low-molecular-weight material was removed by biochemical fractionation. The resulting homogeneous LcrH bound protein A-V antigen fusion peptide but not protein A in a sandwich enzyme-linked immunosorbent assay, and this reaction was inhibited by V(h). These observations indicate that LcrH normally binds V antigen in bacterial cytoplasm and suggest that only free LcrH down-regulates expression of the low calcium response.

Original languageEnglish (US)
Pages (from-to)4313-4318
Number of pages6
JournalInfection and Immunity
Volume64
Issue number10
StatePublished - 1996
Externally publishedYes

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Active Immunity
Plague
Staphylococcal Protein A
Calcium
Antigens
Peptides
Molecular Weight
Bacterial Antigens
Yersinia pestis
Lethal Dose 50
Guanidine
Operon
Affinity Chromatography
Urea
Cytoplasm
Vaccination
Plasmids
Down-Regulation
Enzyme-Linked Immunosorbent Assay
Cytokines

ASJC Scopus subject areas

  • Immunology

Cite this

V antigen-polyhistidine fusion peptide : Binding to LcrH and active immunity against plague. / Motin, Vladimir; Nedialkov, Yuri A.; Brubaker, Robert R.

In: Infection and Immunity, Vol. 64, No. 10, 1996, p. 4313-4318.

Research output: Contribution to journalArticle

Motin, Vladimir ; Nedialkov, Yuri A. ; Brubaker, Robert R. / V antigen-polyhistidine fusion peptide : Binding to LcrH and active immunity against plague. In: Infection and Immunity. 1996 ; Vol. 64, No. 10. pp. 4313-4318.
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