Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 1999 Aug;67(8):3793-9.
doi: 10.1128/IAI.67.8.3793-3799.1999.

Analysis of the immunological responses to transferrin and lactoferrin receptor proteins from Moraxella catarrhalis

R H Yu  1 , R A Bonnah, S Ainsworth, A B Schryvers
Affiliations

Analysis of the immunological responses to transferrin and lactoferrin receptor proteins from Moraxella catarrhalis

R H Yu et al. Infect Immun. 1999 Aug.

Abstract

Moraxella catarrhalis expresses surface receptor proteins that specifically bind host transferrin (Tf) and lactoferrin (Lf) in the first step of the iron acquisition pathway. Acute- and convalescent-phase antisera from a series of patients with M. catarrhalis pulmonary infections were tested against Tf and Lf receptor proteins purified from the corresponding isolates. After the purified proteins had been separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting, we observed strong reactivity against Tf-binding protein B (TbpB; also called OMP1) and Lf-binding protein B (LbpB) but little or no reactivity against Tf-binding protein A (TbpA) or Lf-binding protein A (LbpA), using the convalescent-phase antisera. Considerable antigenic heterogeneity was observed when TbpBs and LbpBs isolated from different strains were tested with the convalescent-phase antisera. Comparison to the reactivity against electroblotted total cellular proteins revealed that the immune response against LbpB and TbpB constitutes a significant portion of the total detectable immune response to M. catarrhalis proteins. Preparations of affinity-isolated TbpA and LbpA reacted with convalescent-phase antisera in a solid-phase binding assay, but blocking with soluble TbpB, soluble LbpB, or extracts from an LbpA(-) mutant demonstrated that this reactivity was attributed to contaminants in the TbpA and LbpA preparations. These studies demonstrate the immunogenicity of M. catarrhalis TbpB and LbpB in humans and support their potential as vaccine candidates.

PubMed Disclaimer

Figures

FIG. 1
FIG. 1
Assessment of the ability of Tf and Lf receptor proteins to elicit an immune response. Using hTf-Sepharose (A) or hLf-Sepharose (B) as the affinity matrix, we isolated receptor proteins from iron-starved cells of M. catarrhalis 4223 and N141. The proteins were eluted by boiling in Laemmli sample buffer and subjected to SDS-PAGE (7% gel), and Western blots were prepared. The blots were either stained for protein (lanes 1, 2) or blocked and then probed with rabbit polyclonal antiserum (lanes 3 and 4; 1/5,000 dilution), human convalescent-phase serum (lanes 5 and 6; 1/200 dilution), or human acute-phase serum (lanes 7 and 8; 1/200 dilution). The human sera were obtained from the patient with a pulmonary infection attributed to M. catarrhalis N141.
FIG. 2
FIG. 2
Analysis of host immune response to Tf and Lf receptor proteins from homologous and heterologous M. catarrhalis strains. Using hTf-Sepharose or hLf-Sepharose as the affinity matrix, we isolated receptor proteins from iron-starved cells of M. catarrhalis 4223, Q8, and N141. The proteins were eluted from the matrix by boiling in Laemmli sample buffer and subjected to SDS-PAGE (7% gel), and replicate Western blots were prepared. The blots were either stained for protein (A) or blocked and probed with convalescent-phase antisera (1/200 dilution) obtained from patients infected with M. catarrhalis N137 (B), N141 (C) or N132 (D).
FIG. 3
FIG. 3
Reactivity of convalescent-phase sera to recombinant Tf and Lf receptors. The regions encoding the mature forms of the individual Tf and Lf receptor proteins were PCR amplified and ligated in frame with the malE gene of the pMal-c2 vector for production of Mbp fusion proteins. Expression of the Mbp fusions was induced by the addition of IPTG, and the cells were boiled in Laemmli sample buffer prior to SDS-PAGE (7% gel). The electroblotted proteins were either stained for protein (lanes 1 to 4) or probed with convalescent-phase antiserum (1:500) from the patient with a pulmonary infection attributed to M. catarrhalis N141. Lanes 1 and 5, Mbp::TbpA (strain 4223 tbpA gene); lanes 2 and 6, Mbp::TbpB (strain 4223 tbpB gene); lanes 3 and 7, Mbp::LbpA (strain N141 lbpA gene); lanes 4 and 8, Mbp::LbpB (strain N141 lbpB gene). Symbols are as in Fig. 1 and 2.
FIG. 4
FIG. 4
Analysis of immune response to M. catarrhalis total cellular proteins. The Tf receptor proteins from M. catarrhalis 4223 (lanes 2, 5, 8, and 11) or the Lf receptors from strain N141 (lanes 3, 6, 9, and 12) were isolated by using either hTf-Sepharose or hLf-Sepharose as the affinity matrix. The receptor proteins or strain N141 whole cells (lanes 1, 4, 7, and 10) were boiled in Laemmli sample buffer and subjected to SDS-PAGE (7% gel), and replicate Western blots were prepared. The blots were either stained for protein (lanes 1 to 3) or blocked and probed with either acute-phase sera from the patient infected with M. catarrhalis N141 (lanes 4 to 6), the convalescent-phase sera alone (lanes 7 to 9), or the convalescent-phase sera containing approximately 1 mg each of Mbp::TbpB and Mbp::LbpB purified by amylose affinity chromatography.
FIG. 5
FIG. 5
Solid-phase immunological analysis. Native Tf receptor proteins were affinity isolated with apo- or holo-hTf-Sepharose from M. catarrhalis 4223 (TbpA and TbpB, TbpA alone, or TbpB alone). Native Lf receptor proteins were affinity isolated with holo-hLf-Sepharose from strain N141 or from an N141 LbpB− isogenic mutant (9). The Mbp receptor protein fusions were affinity isolated by using an amylose resin. Approximately 50 ng of isolated protein was spotted onto the nitrocellulose matrix and incubated with a 1:1,000 dilution of convalescent-phase sera from the patient infected with M. catarrhalis N141 (sera alone; columns 1 and 4), the same sera preincubated with approximately 1 mg of either Mbp::TbpB (column 2) or Mbp::LbpB (column 3) or with 100 μl of a crude extract from the N141 LbpA− isogenic mutant (9) (column 5).
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Aebi C, Cope L D, Latimer J L, Thomas S E, Slaughter C A, McCracken G H, Jr, Hansen E J. Mapping of a protective epitope of the CopB outer membrane protein of Moraxella catarrhalis. Infect Immun. 1998;66:540–548. - PMC - PubMed
    1. Aebi C, Maciver I, Latimer J L, Cope L D, Stevens M K, Thomas S E, McCracken G H, Jr, Hansen E J. A protective epitope of Moraxella catarrhalis is encoded by two different genes. Infect Immun. 1997;65:4367–4377. - PMC - PubMed
    1. Aebi C, Stone B, Beucher M, Cope L D, Maciver I, Thomas S E, McCracken G H, Jr, Sparling P F, Hansen E J. Expression of the CopB outer membrane protein by Moraxella catarrhalis is regulated by iron and affects iron acquisition from transferrin and lactoferrin. Infect Immun. 1996;64:2024–2030. - PMC - PubMed
    1. Ala’Aldeen D A, Davies H A, Wall R A, Borriello S P. The 70 kilodalton iron regulated protein of Neisseria meningitidis is not the human transferrin receptor. FEMS Microbiol Lett. 1990;69:37–42. - PubMed
    1. Ala’Aldeen D A A, Borriello S P. The meningococcal transferrin-binding proteins 1 and 2 are both surface exposed and generate bactericidal antibodies capable of killing homologous and heterologous strains. Vaccine. 1996;14:49–53. - PubMed

Publication types

MeSH terms