Targeting bacterial transferrin and lactoferrin receptors for vaccines
Anthony B. Schryvers
Abstract
The presence of transferrin (Tf) receptors in Gram-negative bacteria that reside in the upper respiratory tract of mammals and birds indicate that they have existed for over 320 million years.Tf/lactoferrin receptors are essential for survival of Gram-negative bacterial pathogens (pathobionts) on the mucosal surface and during invasive infection and thus are ideal targets for vaccines.Structure-based antigen engineering can improve the immune response induced by Tf receptor proteins.Proof of concept experiments in pigs demonstrate that antigens targeting Tf receptors can prevent infection and colonization. A substantial disease burden in vertebrates is due to Gram-negative bacteria that exclusively inhabit the upper respiratory or genitourinary tracts of their hosts and rely on directly acquiring iron from the host iron-binding glycoproteins through surface receptor proteins. The receptors enable these bacteria to proliferate independently from their neighbors on the mucosal surface and during invasive infection of the host. The diversity in these receptors evolved over millions of years of evolution, which thus bodes well for long-lasting vaccine coverage. Experiments in food production animals provide proof of concept for the use of engineered antigens derived from the receptor proteins to prevent colonization and invasive infection in the natural host, strongly supporting development of these vaccines for use in humans. A substantial disease burden in vertebrates is due to Gram-negative bacteria that exclusively inhabit the upper respiratory or genitourinary tracts of their hosts and rely on directly acquiring iron from the host iron-binding glycoproteins through surface receptor proteins. The receptors enable these bacteria to proliferate independently from their neighbors on the mucosal surface and during invasive infection of the host. The diversity in these receptors evolved over millions of years of evolution, which thus bodes well for long-lasting vaccine coverage. Experiments in food production animals provide proof of concept for the use of engineered antigens derived from the receptor proteins to prevent colonization and invasive infection in the natural host, strongly supporting development of these vaccines for use in humans. A number of important Gram-negative pathogens of humans and food production animals that are restricted to a few bacterial lineages (Table 1) possess surface receptors for acquiring iron from the host glycoprotein transferrin (Tf) (see Glossary) and, in some lineages, lactoferrin (Lf) (Figure 1) [1.Ostan N. et al.A comparative, cross-species investigation of the properties and roles of transferrin- and lactoferrin-binding protein B from pathogenic bacteria.Biochem. Cell Biol. 2017; 95: 5-11Crossref PubMed Scopus (6) Google Scholar]. The two-component receptors are comprised of a surface lipoprotein, Tf-binding protein B (TbpB) or Lf-binding protein B (LbpB), that captures iron-loaded Tf or Lf and delivers it to the TonB-dependent outer membrane receptors, Tf-binding protein A (TbpA) or Lf-binding protein A (LbpA). TbpB and LbpB are comprised of two lobes (structurally equivalent domains of a protein) with the N-lobe binding to the iron-loaded C-lobe of Tf or Lf. The demonstration that a portion of the N‐terminal anchoring peptide is required for interacting with TbpA [2.Yang X. et al.The anchor peptide of transferrin binding protein B is required for interaction with transferrin binding protein A.J. Biol. Chem. 2011; 286: 45165-45173Abstract Full Text Full Text PDF PubMed Scopus (21) Google Scholar] and that portions of the anchor peptide that normally wrap around the TbpB C-lobe can assume an alpha-helical structure [3.Calmettes C. et al.Structural variations within the transferrin binding site on transferrin binding protein, TbpB.J. Biol. Chem. 2011; 286: 12683-12692Abstract Full Text Full Text PDF PubMed Scopus (39) Google Scholar] provides insights into the mechanism by which TbpB delivers Tf to TbpA. TbpA or LbpA extract the iron from Tf or Lf and transport it across the outer membrane. As with other TonB-dependent transporters (TBDTs), TbpA and LbpA consist of an outer beta-barrel embedded in the membrane and an inner plug domain that facilitates the transport of the metal ion across the outer membrane using energy derived from an inner membrane TonB complex. The iron is transferred to a periplasmic iron-binding protein (FbpA) that shuttles the iron across the periplasmic space to an inner membrane transport complex.Table 1Bacterial pathogens of humans and food production animals with Tf and Lf receptorsHostPathogenDiseaseTf receptorLf receptorLineageHumanNeisseria meningitidisMeningitis, sepsisYesaYes indicates presence of the bipartite TbpB–TbpA receptor, TbpA2 indicates the single-component receptor.YesaYes indicates presence of the bipartite TbpB–TbpA receptor, TbpA2 indicates the single-component receptor.NeisseriaceaeNeisseria gonorrhoeaeGonorrheaYesYesNeisseriaceaeHaemophilus influenzae(type b)Meningitis, sepsis, pneumoniaYesPasteurellaceaeH. influenzae(non-typeable)Otitis media, chronic obstructive pulmonary disease (COPD), pneumoniaYesPasteurellaceaeMoraxella catarrhalisOtitis media, COPDYesYesMoraxellaceaeCattleMannheimia haemolyticaBovine respiratory disease (BRD)YesPasteurellaceaePasteurella multocidaHemorrhagic septicemia (HS), BRDTbpA2PasteurellaceaeHistophilus somniBRD, thrombotic meningoencephalitis (TME), myocarditisYesTbpA2PasteurellaceaeMoraxella bovisPinkeye (infectious bovine keratoconjunctivitis) IBKYesYesMoraxellaceaeSheepBibersteinia trehalosiSepticemia, pneumoniaYesPasteurellaceaePigActinobacillus pleuropneumoniaePleuropneumoniaYesPasteurellaceaeHaemophilus parasuisGlasser’s diseaseYesPasteurellaceaeActinobacillus suisPneumonia, septicemiaYesPasteurellaceaePoultryAvibacterium sp.NoneYesPasteurellaceaea Yes indicates presence of the bipartite TbpB–TbpA receptor, TbpA2 indicates the single-component receptor. Open table in a new tab An appreciation of the critical role that the Tf and Lf receptors play in the survival of bacteria and their proliferation on the mucosal surface is best acquired by reflecting on how Tf and Lf receptors ultimately arose and evolved as part of an ongoing adaptation to iron availability during development of life on earth [4.Ostan N.K.H. et al.Lactoferrin receptors in Gram-negative bacteria: an evolutionary perspective.Biochem. Cell Biol. 2021; 99: 102-108Crossref PubMed Scopus (12) Google Scholar]. Possession of these receptor proteins enable the bacteria to proliferate independently from other members of the microbial community and cause localized or systemic infections, making the receptor proteins ideal targets for vaccines. The importance of iron for life on earth is thought to be a consequence of there being high levels of ferrous iron in primordial seas, which facilitated its use as a catalyst for biological oxidation–reduction reactions. The progressive oxygenation of the Earth’s atmosphere and oceans, with key stages at 2.3 billion and 800 million years ago [5.Parnell J. et al.Early oxygenation of the terrestrial environment during the Mesoproterozoic.Nature. 2010; 468: 290-293Crossref PubMed Scopus (80) Google Scholar], impacted the solubility of the ferric ion and created the need for development of systems for acquiring iron under limiting conditions. This led to production and secretion of iron-binding molecules of increasing affinity (ancestral siderophores) by microorganisms, along with specialized systems for binding and uptake, resulting in iron-containing complexes. In Gram-negative bacteria, this likely consisted of ancestral TBDTs such as siderophore receptors with a cognate periplasm to cytoplasm transport pathway (Figure 2, left panel). There is evidence for microbial communities (stromatolites) that characteristically contain cyanobacteria (photosynthetic Gram-negative prokaryotes) being present in geological formations as early as 3.5 billion years ago [6.Rishworth G.M. et al.Non-reliance of metazoans on stromatolite-forming microbial mats as a food resource.Sci. Rep. 2017; 7: 42614Crossref PubMed Scopus (25) Google Scholar]. This suggests that the ubiquitous presence of microbial communities (biofilms) in virtually all ecological niches today was likely established early in evolutionary history. TBDTs for other metal ions such as zinc also arose early in the evolution of Gram-negative bacteria and have subsequently expanded to include TBDTs directed against host proteins that sequester the metal ion [7.Lonergan Z.R. Skaar E.P. Nutrient zinc at the host-pathogen interface.Trends Biochem. Sci. 2019; 44: 1041-1056Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar]. However, zinc acquisition has not involved anything equivalent to siderophore production for iron (Figure 2, left panel). The development of an extensive spectrum of siderophores was primarily thought to be driven by competition between bacteria, but they clearly are part of the complex interactions within microbial communities [8.Kramer J. et al.Bacterial siderophores in community and host interactions.Nat. Rev. Microbiol. 2019; 18: 152-163Crossref PubMed Scopus (337) Google Scholar], such as in modern day sea sediment, where there are a limited number of siderophore producers and many neighboring bacteria dependent on the production of siderophores [9.D'Onofrio A. et al.Siderophores from neighboring organisms promote the growth of uncultured bacteria.Chem. Biol. 2010; 17: 254-264Abstract Full Text Full Text PDF PubMed Scopus (327) Google Scholar]. Single-lobed iron-binding proteins ancestral to bi-lobed Tf arose in early metazoans to acquire iron from the aqueous environment and likely to also compete with the ubiquitous microbial communities. Bi-lobed iron-binding proteins arose by gene duplication events in various metazoan lineages [10.Hernandez-Pasos J. et al.Melanotransferrin: new homolog genes and their differential expression during intestinal regeneration in the sea cucumber Holothuria glaberrima.J. Exp. Zool. B Mol. Dev. Evol. 2017; 328: 259-274Crossref PubMed Scopus (12) Google Scholar], including ancestral serum Tf, at around 670 million years ago [11.Lambert L.A. Molecular evolution of the transferrin family and associated receptors.Biochim. Biophys. Acta. 2012; 1820: 244-255Crossref PubMed Scopus (95) Google Scholar]. The transition to life on land about 420 million years ago clearly impacted the microbial community on mucosal surfaces of vertebrates and led to ancestral Tf being present on mucosal surfaces. The evolutionary process for development of Tf receptors likely initially involved a single-component receptor (Figure 2, middle panel) arising from an ancestral TBDT that transported ferric iron and ultimately being replaced by a more efficient two-component receptor (Figure 2, right panel) that arose by around 320 million years ago [4.Ostan N.K.H. et al.Lactoferrin receptors in Gram-negative bacteria: an evolutionary perspective.Biochem. Cell Biol. 2021; 99: 102-108Crossref PubMed Scopus (12) Google Scholar]. Tf has iron to its C-lobe under and is the by the bipartite receptor (Figure 2, right the ancestral single-component receptor also have the C-lobe of Tf (Figure 2, middle for Tf there are systems of the stages that have involved in to the ancestral bipartite receptor a TbpB it strongly for the importance of the bi-lobed TbpB in the of iron acquisition from Tf under in the upper respiratory tract but evidence is The TbpA2 present exclusively in modern day Gram-negative bacteria that the or of of a transferrin receptor in bovine of PubMed Scopus Google A. of and and a homolog in and of iron availability on their PubMed Scopus Google A. et two systems for acquisition of PubMed Scopus Google Scholar] (Figure 2, middle for Tf likely arose in response to the on the in million years The TbpA2 in and to the N-lobe (Figure 2, middle for Tf in to the bipartite receptor present in the pathogens and (Table 1) that to the C-lobe of Tf for Tf In of the in binding to bovine Tf, and the in the and of the of the TbpA2 has some to the and in the plug of and and of bacterial transferrin and lactoferrin receptor protein PubMed Scopus Google Scholar] that are in of this is a of evolution in which transport of an ferric ion by a TBDT a There is an pathway for iron from the outer membrane into the cytoplasm (Figure 2, middle panel) in of from a of hosts that has high with the pathway from for bovine of possess the TbpA2 receptor, there clearly are other TBDTs in of that are of ferric ion acquired from other and likely the of TBDTs that to the Tf and Lf of the on the surface of indicate that involved in binding to the bacterial receptor protein TbpA and ultimately for the host within over a from bacterial iron through evolution of PubMed Scopus Google Scholar]. of this to the of receptors in A. et al.Bacterial receptors for host transferrin and and role in Microbiol. PubMed Scopus Google Scholar] and between the of to acquire iron and the expression of PubMed Scopus (25) Google Scholar] indicates that Tf has on mucosal surfaces for at 320 million the ancestral lineages to birds and mammals [4.Ostan N.K.H. et al.Lactoferrin receptors in Gram-negative bacteria: an evolutionary perspective.Biochem. Cell Biol. 2021; 99: 102-108Crossref PubMed Scopus (12) Google Scholar]. An early of the between and receptor that be with Tf affinity was to the and and of bacterial transferrin and lactoferrin receptor protein PubMed Scopus Google Scholar] to a from from bacteria that inhabit hosts but not from bacteria that inhabit there is between and and between and likely a of the of evolution from the ancestral proteins. Lf arose in the by gene duplication that between and million years ago [11.Lambert L.A. Molecular evolution of the transferrin family and associated receptors.Biochim. Biophys. Acta. 2012; 1820: 244-255Crossref PubMed Scopus (95) Google et of lactoferrin promote in and modern PubMed Scopus Google Scholar], which it to acquire new and as it was not required for the critical role in iron was for its important role of limiting iron availability to in and at of but the of to this protein has expanded a Cell Biol. 2012; PubMed Scopus Google Scholar]. the Lf receptor genes have from Tf receptor genes Lf was as a iron on the mucosal surface of the upper respiratory but it is that it or was acquired by in two of the bacterial lineages (Table The best the importance of the Tf and Lf receptors for survival on the mucosal surface of the host from experiments with in et infection in with and Microbiol. 2011; PubMed Scopus Google Scholar], that receptors are not of infection and that the Tf receptor is more at supporting infection the Lf receptor. levels as a for mucosal it was that Tf was at levels Lf the bacterial an response was for the of the that Lf from Tf is the presence of a N‐terminal that can in the of a Cell Biol. 2012; PubMed Scopus Google Scholar], which likely was the for the acquisition of in in of the LbpB C-lobe A. et al.The of lactoferrin binding protein an adaptation against PubMed Scopus Google Scholar]. have to against and consist of as as in the bovine pathogens that cause The receptors are present in a few bacterial lineages A. et al.Bacterial receptors for host transferrin and and role in Microbiol. 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