Distinct roles of the N-terminal and C-terminal precursor domains in the biogenesis of the Bordetella pertussis filamentous hemagglutinin

Abstract

The 220-kDa Bordetella pertussis filamentous hemagglutinin (FHA) is the major exported protein found in culture supernatants. The structural gene of FHA has a coding potential for a 367-kDa protein, and the mature form constitutes the N-terminal 60% of the 367-kDa precursor. The C-terminal domain of the precursor was found to be important for the high-level secretion of full-length FHA but not of truncated analogs ( Exported proteins in gram-negative microorganisms face the challenge of having to cross two distinct membranes, called the inner and outer membranes. In most cases, translocation through the inner membrane involves the signal peptide-dependent general secretion pathway (for a recent review, see reference 22). Important exceptions include the members of protein families related to the RTX hemolysins (31). Transport across the outer membrane often requires the concerted action of several accessory proteins that were thought to be specific for a given exported protein but that appear now to fall within distinct protein families.Successful interaction of bacterial pathogens with their hosts usually requires the production of several extracellular proteins that constitute important virulence factors. Therefore, pathogenic organisms represent interesting models for the study of protein export, especially across the outer membrane. Bordetella pertussis, the etiologic agent of whooping cough, produces several virulence factors that are located at the outer surface of the organism or released into the extracellular milieu. These factors include pertussis toxin, adenylate cyclase toxin, filamentous hemagglutinin (FHA), fimbriae, and probably other proteins that have yet to be identified. Accessory genes important for the biogenesis and export of several of these factors have been isolated and studied (6,14,30,32,33). Interestingly, in each case the accessory proteins were found to be members of protein families involved in export and biogenesis of macromolecules in other bacterial genera.Among the various protein export systems of B. pertussis, the FHA export machinery is particularly efficient, because FHA represents the major secreted protein in B. pertussis culture supernatants. This high efficiency is particularly interesting because of the large size of monomeric FHA polypeptides (220 kDa).FHA is a major adhesin produced by several Bordetella species. It expresses at least three different adherence activities (for a recent review, see reference 13). The region downstream of the FHA structural gene (fhaB) contains several open reading frames, of which the three most proximal are involved in the biogenesis of the fimbriae (14,32,33) and the most distal is involved in the biogenesis of FHA (32). The protein encoded by this latter gene, named fhaC, has significant sequence similarities with accessory proteins involved in the secretion of hemolysins of Serratia marcescens (20) and Proteus mirabilis (27). In addition, the N-terminal region of FHA has strong sequence similarities with the N-terminal domains of t...

Bordetellafilamentous hemagglutinin plays a critical role in immunomodulation, suggesting a mechanism for host specificity

“…A third possibility is that FHA Bp is inherently immunomodulatory (even in rodents), but this ability requires that the bacteria first be able to adhere specifically to host cells, i.e., that adherence and immunomodulation are mediated by separate domains within FHA and that each domain may display different abilities to distinguish receptors from different hosts. This hypothesis is supported by the fact that multiple binding domains within FHA Bp , with apparently different specificity, have been identified (20,29,30,(55)(56)(57).…”

Lack of functional complementation between Bordetella pertussis filamentous hemagglutinin and Proteus mirabilis HpmA hemolysin secretion machineries

“…This highly conserved segment is hypothesized to interact with the cognate accessory protein. In agreement with this proposal, the deletion of this region totally abolishes the secretion of FHA (35), whereas several truncated forms of FHA which retain this region are secreted very efficiently in a FhaC-dependent manner (28). Small in-frame deletions as well as certain point mutations within this segment of ShlA affected both its secretion and activation (31).…”

“…This N-terminal maturation appears to be much more extensive for HMWA than for FHA. In addition and unlike that of the hemolysins, the secretion of FHA involves extensive C-terminal processing of a large precursor (10,27,28). Finally, the hemolysins acquire hemolytic activity via activation by their cognate accessory proteins, whereas the adhesins are unlikely to require activation.…”

The crystal structure of filamentous hemagglutinin secretion domain and its implications for the two-partner secretion pathway

“…A 5Ј segment of fhaB was amplified by PCR with the oligonucleotides 5Ј-ATGAACAC-GAACCTGTACAGG-3Ј and 5Ј-GGTACCTCAGGATCCCT-TGCCGCCGCCTTGCA-3Ј. The amplicon was cloned in pC-RII-Topo (Invitrogen) and sequenced by using the ABI 377 Prism sequencer, and the 1.1-kb SphI-KpnI fragment of the recombinant plasmid was exchanged for the 1.3-kb SphI-KpnI fragment of pBG12 (15), yielding pFJD95. The EcoRI-KpnI fragment of pFJD95, corresponding to the first 1.4 kb of fhaB, was then introduced into pUC19, generating pFJD117.…”