Analysis of a Unique Interaction between the Complement Regulatory Protein Factor H and the Periodontal PathogenTreponema denticola

Abstract

Treponema denticola, a spirochete associated with periodontitis, is abundant at the leading edge of subgingival plaque, where it interacts with gingival epithelia. T. denticola produces a number of virulence factors, including dentilisin, a protease which is cytopathic to host cells, and FhbB, a unique T. denticola lipoprotein that binds complement regulatory proteins. Earlier analyses suggested that FhbB specifically bound to factor H (FH)-like protein 1 (FHL-1). However, by using dentilisin-deficient mutants of T. denticola, we found that T. denticola preferentially binds FH and not FHL-1, and that FH is then cleaved by dentilisin to yield an FH subfragment of ϳ50 kDa. FH bound to dentilisin-deficient mutants but was not cleaved and retained its ability to serve as a cofactor for factor I in the cleavage of C3b. To assess the molecular basis of the interaction of FhbB with FH, mutational analyses were conducted. Replacement of specific residues in widely separated domains of FhbB and disruption of a central alpha helix with coiled-coil formation probability attenuated or eliminated FH binding. The data presented here are the first to demonstrate the retention at the cell surface of a proteolytic cleavage product of FH. The precise role of this FH fragment in the host-pathogen interaction remains to be determined.Adult periodontitis, the most common infection of middleaged adults, affects approximately 116 million adults in the United States (3). Periodontal disease is a multifactorial process involving alterations of the overall composition of the oral flora coupled with host-determined susceptibility factors (73). The process is initiated by the formation and spread of polymicrobial biofilms that ultimately progress to plaque formation. The human oral cavity may contain up to 700 bacterial species (58), with the subgingival plaque estimated to consist of up to 415 species (57). Treponema denticola is one of the dominant spirochete species at the leading edge of plaque, and a clear correlation has been demonstrated between its abundance and the occurrence and severity of periodontal disease (reviewed in reference 15).We previously reported that T. denticola binds at least one member of the factor H (FH) protein family (43). In humans, the FH protein family consists of FH, FH-like protein 1 (FHL-1), and five FH-related proteins designated FHR1 through FHR5 (35). FH and FHL-1 serve as cofactors in the factor I-mediated cleavage of C3b, a key opsonin, and accelerate the decay of the C3 convertase complex, leading to downregulation of C3b production (56,66,67,74). Evasion of complement by oral bacteria such as T. denticola is essential as complement proteins are present in gingival fluid at levels as high as 85% of that reported for serum (68). In addition, there is evidence that complement is more active in saliva than in serum (8, 9). The binding to cell-anchored FH family proteins by some microbial pathogens has also been demonstrated to be an important adherence-and-invasion mechanism (5, 23, 55).The interacti...

Structure of Factor H-binding Protein B (FhbB) of the Periopathogen, Treponema denticola

“…Substitution analyses indicate that intramolecular hydrogen bonds between Asn-44 -Lys-47 (␣1-turn 1), Glu-45-Arg-49 (␣1-turn 1), and His-50 -Gln-85 (turn 1-turn 2) are important for FH binding. Other FhbB residues that have been demonstrated to influence FH binding include Ile-63, Leu-68, Phe-96, and the C-terminal seven amino acids (23). The placement of these residues on the FhbB structure suggests that their influence on FH binding is indirect as they are not surfaceexposed.…”

“…Loss of FH binding by an I63T substitution mutant most likely results from aberrant hydrophobic packing within ␣2. A Phe-96 substitution mutant and C-terminal 7-amino acid truncation mutant have also been shown to lack FH-binding ability (23). The structure of FhbB indicates that several C-terminal residues, including Phe-96, reside within the hydrophobic core of the protein forming a two-stranded ␤-sheet.…”

“…FhbB is the smallest (11.4 kDa) bacterially produced FH-binding protein identified to date (20,21). FhbB binds and positions FH on the cell surface thus allowing FH cleavage by the T. denticola protease, dentilisin (22)(23)(24). It is our hypothesis that in vivo, FH cleavage leads to its depletion in the subgingival crevice resulting in local dysregulation of complement and conditions that favor the development and progression of periodontal disease.…”

Treponema denticola chymotrypsin-like proteinase (CTLP) integrates spirochaetes within oral microbial communities

“…One of these is a surface-localized prolyl-phenylalanine-specific chymotrypsin-like protease (CTLP) with a broad repertoire of cytopathic activities (Fenno et al, 1998b;Ellen et al, 2000). The CTLP, also designated dentilisin (Ishihara et al, 1996), has been reported to degrade a range of host proteins (Bamford et al, 2007;McDowell et al, 2009), hydrolyse bioactive peptides (Mäkinen et al, 1995;Miyamoto et al, 2006) and help spirochaetes penetrate epithelial cell monolayers (Fenno et al, 1998b;Ellen et al, 2000;Chi et al, 2003). Formation of the CTLP functional complex on the cell surface requires expression of three genes, prcB, prcA and prtP (Godovikova et al, 2010).…”

Gene Regulation, Two Component Regulatory Systems, and Adaptive Responses in Treponema Denticola

“…Important to both metabolism and virulence, T. denticola strains encode an array of 18-25 proteases. Two proteases that have been experimentally characterized in some detail are dentilisin (Chi et al 2003;Ellen et al 2000;Ishihara et al 1998;McDowell et al 2009;Miller et al 2016;Yamazaki et al 2006) and cystalysin (Chu et al 2002(Chu et al , 2008Krupka et al 2000;Zhang et al 2010). Both play important roles in T. denticola pathogenesis.…”