A Novel Fibronectin Binding Motif in MSCRAMMs Targets F3 Modules

Abstract

BackgroundBBK32 is a surface expressed lipoprotein and fibronectin (Fn)-binding microbial surface component recognizing adhesive matrix molecule (MSCRAMM) of Borrelia burgdorferi, the causative agent of Lyme disease. Previous studies from our group showed that BBK32 is a virulence factor in experimental Lyme disease and located the Fn-binding region to residues 21–205 of the lipoprotein.Methodology/Principal FindingsStudies aimed at identifying interacting sites between BBK32 and Fn revealed an interaction between the MSCRAMM and the Fn F3 modules. Further analysis of this interaction showed that BBK32 can cause the aggregation of human plasma Fn in a similar concentration-dependent manner to that of anastellin, the superfibronectin (sFn) inducing agent. The resulting Fn aggregates are conformationally distinct from plasma Fn as indicated by a change in available thermolysin cleavage sites. Recombinant BBK32 and anastellin affect the structure of Fn matrices formed by cultured fibroblasts and inhibit endothelial cell proliferation similarly. Within BBK32, we have located the sFn-forming activity to a region between residues 160 and 175 which contains two sequence motifs that are also found in anastellin. Synthetic peptides mimicking these motifs induce Fn aggregation, whereas a peptide with a scrambled sequence motif was inactive, suggesting that these motifs represent the sFn-inducing sequence.Conclusions/SignificanceWe conclude that BBK32 induces the formation of Fn aggregates that are indistinguishable from those formed by anastellin. The results of this study provide evidence for how bacteria can target host proteins to manipulate host cell activities.

Borrelia burgdorferi Protein BBK32 Binds to Soluble Fibronectin via the N-terminal 70-kDa Region, Causing Fibronectin to Undergo Conformational Extension

“…However, there is evidence that a sequence within the same region of BBK32, underlined in Fig. 1B, binds to isolated FNIII modules (24) and, as observed for the anastellin fragment derived from 1 FNIII of FN (25), induces FN aggregation. Here, we have employed complementary techniques to characterize the interaction of Bbk32 with FN.…”

Fibronectin-binding nanoparticles for intracellular targeting addressed by B. burgdorferi BBK32 protein fragments

“…Different BBK32 peptides used in this study have been shown to bind different FN regions. [25][26][27][28] Our results show that only BF130-166 VLPs demonstrated clear, strong FN-binding ability. Moreover, we were able to show that this binding occurs via GBD of FN.…”

“…However, induction of ordered aggregation of soluble FN and inhibition of endothelial cell proliferation similar to that of anastellin were shown for these BBK32 fragments, so there may be further interest in these constructs in the future. 27 Recently, retroviral binding to FN was associated with increased infection of target cells and efficient gene transfer. 21,38 Therefore, we tested the possible intracellular entry of the BF130-166 VLPs using BHK-21 as target cells.…”

“…25 It was shown that the region of BBK32 comprising amino acids 147 -205 binds to the N-terminal FNI and FNIII modules, but residues 120 -147 of BBK32 protein are considered similar to the FN-binding region of SfbI protein from S. pyogenes that had been shown to bind to the GBD of FN. [26][27][28] Here we present the construction of HBc-based chimeric VLPs with FN-binding property by insertion of B. burgdorferi BBK32 fragments into the MIR region of capsid. The evidence of intracellular internalization of these VLPs into nonphagocytic cells through caveolae-mediated endocytosis makes this construct an attractive model in development of HBc-based NPs for medically relevant cellular targeting applications and highlights the 130 -166 aa fragment of B. burgdorferi BBK32 as a novel cellular uptake-promoting peptide.…”

Conformational remodeling of the fibronectin matrix selectively regulates VEGF signaling

“…The cryptic polymerization activity present in the III 1 domain of fibronectin can be recapitulated in a peptide termed anastellin (Morla et al, 1994;Morla and Ruoslahti, 1992). Subsequent studies showed that anastellin can bind directly to previously assembled fibronectin matrix and alter its conformation (Klein et al, 2003;Prabhakaran et al, 2009). Anastellin has been found to inhibit tumor angiogenesis in vivo as well as endothelial cell proliferation in vitro; however, its mechanism of action has remained elusive (Ambesi et al, 2005;Yi and Ruoslahti, 2001).…”