Purification of the Neisseria meningitidis transferrin binding protein-2 (TBP2) to homogeneity using column chromatography

Ferrón, L

doi:10.1016/0378-1097(93)90013-r

Cited by 15 publications

(12 citation statements)

References 22 publications

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“…The bacterial transferrin receptors which have been studied are composed of two polypeptide chains: transferrinbinding proteins 1 and 2, Tbp1 and Tbp2, respectively. The transferrin-binding proteins have been purified from strains of H. influenzae (Schryvers, 1989) and N. meningitidis (Danve et al, 1993;Ferron et al, 1993) by affinity chromatography using human transferrin bound to solid supports. In H. influenzae, the molecular mass of Tbp1 is approximately 100 kDa, whereas that of Tbp2 is variable, ranging from 60 to 90 kDa, depending upon the strain (Holland et al, 1992;Schryvers and Gray-Owen, 1992).…”

Section: Introductionmentioning

confidence: 99%

Cloning and expression of the Haemophilus influenzae transferrin receptor genes

Loosmore¹,

Yang²,

Coleman³

et al. 1996

Molecular Microbiology

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The genomic transferrin receptor genes (tbpA and tbpB) from two strains of Haemophilus influenzae type b (Hib) and two strains of non-typable H. influenzae (NTHi) have been cloned and sequenced. The deduced protein sequences of the H. influenzae tbpA genes were 95-100% conserved and those of the tbpB genes were 66-100% conserved. The tbpB gene from one strain of NTHi was found to encode a truncated Tbp2 protein. The tbpB genes from four additional NTHi strains were amplified by the polymerase chain reaction (PCR) utilizing primers derived from the conserved N-terminal sequences of Tbp1 and Tbp2 and were found to encode full-length proteins. Although several bacterial species express transferrin receptors, when the Tbp1 and Tbp2 sequences from different organisms were compared, there was only limited homology. Recombinant Tbp1 and Tbp2 proteins were expressed from Escherichia coli and antisera were raised to the purified proteins. There was significant antigenic conservation of both Tbp1 and Tbp2 amongst H. influenzae strains, as determined by Western blot analysis. In a passive model of bacteraemia, infant rats were protected from challenge with Hib after transfer of anti-rTbp2 antiserum, but not after anti-rTbp1 antiserum.

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Section: Introductionmentioning

confidence: 99%

Cloning and expression of the Haemophilus influenzae transferrin receptor genes

Loosmore¹,

Yang²,

Coleman³

et al. 1996

Molecular Microbiology

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“…However, TbpA alone has shown little promise, with a previous study failing to detect TbpAinduced bactericidal activity (29). Furthermore, antibodies raised against native TbpA or the TbpA plus TbpB complex fail to react with electroblotted TbpA, and purification of active (i.e., hTf binding) protein has been problematic (2,17,29). These findings are related to the difficulties in maintaining the tertiary structure of TbpA when isolated from the membrane (2,36).…”

Section: Discussionmentioning

confidence: 99%

Recombinant Neisseria meningitidis Transferrin Binding Protein A Protects against Experimental Meningococcal Infection

et al. 2001

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To better characterize the vaccine potential of Neisseria meningitidis transferrin binding proteins (Tbps), we have overexpressed TbpA and TbpB from Neisseria meningitidis isolate K454 in Escherichia coli. The ability to bind human transferrin was retained by both recombinant proteins, enabling purification by affinity chromotography. The recombinant Tbps were evaluated individually and in combination in a mouse intraperitoneal-infection model to determine their ability to protect against meningococcal infection and to induce cross-reactive and bactericidal antibodies. For the first time, TbpA was found to afford protection against meningococcal challenge when administered as the sole immunogen. In contrast to the protection conferred by TbpB, this protection extended to a serogroup C isolate and strain B16B6, a serogroup B isolate with a lowermolecular-weight TbpB than that from strain K454. However, serum from a TbpB-immunized rabbit was found to be significantly more bactericidal than that from a TbpA-immunized animal. Our evidence demonstrates that TbpA used as a vaccine antigen may provide protection against a wider range of meningococcal strains than does TbpB alone. This protection appears not to be due to complement-mediated lysis and indicates that serum bactericidal activity may not always be the most appropriate predictor of efficacy for protein-based meningococcal vaccines.

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“…Antisera to strains P000, P391, P636, V002 and V021 were obtained as described previously [9], by hyperimmunization of mice with blebs obtained by culturing in ironrestricted medium and subsequent adsorption of sera with blebs obtained from iron-sufficient medium to eliminate interfering antibodies against lipopolysaceharides and non-IR proteins. Titration of sera was as previously described [13], by a dot-blot procedure in which 2 ~1 aliquots of OMP preparation, deposited onto nitrocellulose membranes, were incubated with five-fold serial dilutions of the corresponding antiserum and with peroxidase-labelled goat IgG raised against human or mouse Igs (POD-IgG). Binding was visualized with chloronaphthol; in all cases titres of 1/125 were obtained, and a working dilution of 1/100 was thus selected.…”

Section: Immune Seramentioning

confidence: 99%

In vivo human immune response to transferrin-binding protein 2 and other iron-regulated proteins of Neisseria meningitidis

Ferreirós¹

1994

FEMS Immunology and Medical Microbiology

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When grown under iron restriction, Neisseria meningitidis expresses new outer-membrane proteins, some of which are antigenic and potentially useful as vaccine components. This is particularly relevant to N. meningitidis serogroup B, against which neither polysaccharide nor conjugate vaccines are effective. We investigated recognition of N. meningitidis serogroup B outer-membrane antigens by three sera from patients recovered from meningitis. Recognition of antigens from the homologous strain provided information on in vivo expression during infection and immunogenicity, while cross-reactivity with outer membrane proteins from the other two strains and from another five strains in our collection allowed evaluation of antigenic heterogeneity. Our results demonstrate that transferrin-binding protein 2 (TBP2) is immunogenic in humans, to varying degrees depending on the strain, and that TBP2s (like the equivalent proteins of Haemophilus influenzae type b) are among the most important iron-regulated outer membrane antigens expressed during infection. Other immunogenic outer membrane proteins (some iron-regulated) are also expressed during infection; in a previous study in mouse, three of these proteins (with M(r) of 50, 70 and 77 kDa) did not induce an immune response. Our cross-reactivity data provide some support for Robki et al.'s two-group classification of N. meningitidis strains, and provide evidence against the possibility that the antigenic domains shared by the TBP2s of all N. meningitidis strains induce immune responses in vivo.

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Purification of the Neisseria meningitidis transferrin binding protein-2 (TBP2) to homogeneity using column chromatography

Cited by 15 publications

References 22 publications

Cloning and expression of the Haemophilus influenzae transferrin receptor genes

Cloning and expression of the Haemophilus influenzae transferrin receptor genes

Recombinant Neisseria meningitidis Transferrin Binding Protein A Protects against Experimental Meningococcal Infection

In vivo human immune response to transferrin-binding protein 2 and other iron-regulated proteins of Neisseria meningitidis

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