Cloning and expression of the gene for a fibronectin-binding protein from Staphylococcus aureus.

Flock, Jan‐Ingmar; Fröman, Gunnar; Jönsson, Klas; Guss, Bengt; Signäs, Christer; Nilsson, Björn; Raucci, Giuseppe; Höök, Magnus; Wadström, Torkel; Lindberg, Martin

doi:10.1002/j.1460-2075.1987.tb02511.x

Cited by 235 publications

(143 citation statements)

References 24 publications

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“…Moreover, a monomodal distribution, like that in Fig. 2a, reappeared after the passivating molecules of Fn were washed out with a diluted solution of acetic acid (25). When a clinically isolated mutant strain (7) that did not express adhesins was used, a distribution of the rupture forces was obtained (Fig.…”

Section: Resultsmentioning

confidence: 83%

Dynamics of the interaction between a fibronectin molecule and a living bacterium under mechanical force

Bustanji

Arciola

Conti

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

103

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C ell-protein adhesion regulates essential processes in multicellular organisms and provides signals that affect the morphology, motility, gene expression, and survival of cells (1). Fibronectin (Fn) is an extracellular matrix protein that is widely distributed in the tissues of all vertebrates and is a potential ligand for most cell types (2). Fn is also an important mediator of bacterial invasions and of persistent infections like that of Staphylococcus epidermidis (3). Until now, S. epidermidis was considered a mere saprophyte, usually harboring on the skin and mucosae. Recently, this organism has been shown to also act as a pathogen, mainly in association with surgical applications of biomaterials (4).Like many other cell-protein adhesions, the binding between Fn and staphylococci takes place under physiological shear rates (5). It has been reported that the high shear stress characteristic of the turbulent flow at blood vessel entrances and bifurcations can prevent Fn-mediated adhesion of Staphylococcus aureus to the endothelial cells lining the vessel, reducing, in this way, the chances of bacterial invasion (6). Thus, a thorough characterization of the binding͞unbinding dynamics of these interactions is necessary to better understand the molecular mechanisms of Fn-mediated pathogenesis. Moreover, specific cell-protein interactions have been selected by nature to mediate different dynamic processes of adhesion of a cell under hydrodynamic flow, as in the case of the rolling adhesion of leukocytes whose velocity in the blood vessels is controlled by transient binding events to the inner walls. It is of interest to understand how different cell͞protein adhesions are optimized in their functions through the details of the interactions at the molecular level. In this article, we address this issue by comparing the interaction dynamics of three different adhesion processes: firm adhesion mediated by the biotin͞avidin pair, rolling adhesion, which is mediated by the L-selectin͞P-selectin glycoprotein ligand 1 (PSGL-1) system, and cell͞bacterium adhesion mediated by Fn͞adhesin binding.Traditionally, adhesion interactions have been studied in reversible equilibrium conditions. However, in vivo, these interactions take place under significant shear forces with pulling rates that are much faster than the relaxation rates of the binding pair, and thus, the binding͞unbinding process occurs under nonequilibrium, irreversible conditions. The development of single-molecule manipulation methods, such as scanning force microscopy (SFM) and optical tweezers, has made it possible to investigate the dynamics of these processes under nonequilibrium conditions, and to measure their force-dependent dissociation kinetics. Here, we investigate the dynamics of the interactions between individual living S. epidermidis cells and single Fn molecules, map the energy landscape of the binding͞ unbinding process, and measure the association and dissociation rate constants of the binding pair by using SFM. In particular, we show that two p...

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Section: Resultsmentioning

confidence: 83%

Dynamics of the interaction between a fibronectin molecule and a living bacterium under mechanical force

Bustanji

Arciola

Conti

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

103

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“…In an earlier report from our laboratories a protein with a Mr Of 18,000 present in lysostaphin lysates of S. aureus was implicated as a FnBP (6 We have earlier reported on the molecular cloning in E. coli of the gene for a FnBP from S. aureus strain 8325-4 (17). In this paper we report the complete nucleotide sequence of the gene.…”

mentioning

confidence: 80%

Nucleotide sequence of the gene for a fibronectin-binding protein from Staphylococcus aureus: use of this peptide sequence in the synthesis of biologically active peptides.

Signäs

Raucci

Jönsson

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

355

295

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Binding of cells of Staphylococcus aureus to fibronectin, which may represent a mechanism of host tissue adherence, involves a fibronectin-receptor protein present on the bacterial surface. Cloning of a gene coding for a staphylococcal fibronectin-binding protein and construction of a fusion protein with fibronectin-binding properties was previously reported from our laboratory. We have now sequenced the gene and deduced a primary sequence of the fibronectin-binding protein. The protein resembles other cell-wall-associated proteins on Gram-positive bacteria in that it (i) appears to be anchored in the cell membrane via its C-terminal end, (ii) contains a proline-rich repeating unit outside the membrane anchor, and (iii) contains a long (36-amino acid) signal sequence at the N terminus. The fibronectin-binding activity has been localized to a domain composed of a 38-amino acid unit repeated completely three times and partially a fourth time; the identity between the three 38-amino acid sequences varies from 42 to 87%. Three synthetic peptides mimicking the structure of each 38-amino acid unit were constructed. All three peptides interacted with fibronectin, as indicated by their ability to inhibit binding of fibronectin to staphylococcal cells, whereas an unrelated 37-amino acid peptide showed no inhibitory activity.

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“…The staphylococcal fusion protein, ZZ-FR, containing two immunoglobu1in-Gbinding domains of protein A and three fibronectin-binding repeats [15], was a gift from G. Raucci (Menarini Ricerche Sud, Italy). Bovine serum albumin, ovalbumin, fetuin, bovine LgG and tyroglobulin were from Sigma Chemical Co., St.…”

Section: Methodsmentioning

confidence: 99%

Multiple binding sites in fibronectin and the staphylococcal fibronectin receptor

Bozzini

Visai

Pignatti

et al. 1992

European Journal of Biochemistry

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The binding of fibronectin to Staphylococci exhibits the properties of a ligand-receptor interaction and has been proposed to mediate bacterial adherence to host tissues.To localize staphylococcal-binding sites in fibronectin, the protein was subjected to limited proteolysis and, of the generated fragments, Staphylococci appeared to preferentially bind to the N-terminal fragment. Different fibronectin fragments were isolated and tested for their ability to inhibit lZ5I-fibronectin binding to Staphylococci. The results indicate that only the N-terminal region effectively competed for fibronectin binding. However, when isolated fragments were adsorbed to microtiter wells, we found that two distinct domains, corresponding to the N-terminal fragment and to the heparin-binding peptide mapping close to the C-terminal end of fibronectin, promoted the attachment of both Staphylococcus aureus Newman and coagulase-negative strain of Staphylococcus capitis 651.These same domains were recognized by purified '251-labeled staphylococcal receptor, either when immobilized on microtiter wells or probed after adsorbtion onto nitrocellulose membrane.The heparin-binding domain is comprised of type-111-homology repeats 14, 15 and 16. To determine which repeats participate in this interaction, we isolated and tested repeats type IIII4 and type IIII6. We found that the major staphylococcal-binding site is located in repeat type III14.The staphylococcal receptor bound the N-terminal domain of fibronectin with a KD of 1.8 nM, whereas the dissociation constant of the receptor molecule for the internal heparin-binding domain was 10 nM.Since the fusion protein ZZ-FR, which contains the active sequences of fibronectin receptor (Dl-D3) bound only to the N-terminus, it is reasonable to assume that the bacterial receptor may have additional binding sites outside the D domains, capable of interacting with the internal heparinbinding domain of fibronectin.Fibronectin is a large dimeric glycoprotein found in body fluids, present in a fibrillar form in the extracellular matrix of loose connective tissue.Fibronectin contains a number of discrete functional domains capable of independent binding to ligands such as collagens, glycosaminoglycans, fibrin(ogen) and thrombospondin. Fibronectin also mediates adhesion of various cell lines through the RGD sequence, located in the central part of the protein. The RGD sequence within fibronectin has been shown to interact with cell-surface glycoproteins, termed integrins, which are a family of structurally related heterodimers. Moreover, the internal heparin-binding domain can also promote cell adhesion by an RGD-independent mechanism. This adhesion can be inhibited by heparin and it is likely that this domain of fibronectin interacts with cellsurface heparan sulfate proteoglycans 111.Several pathogenic Gram-positive bacteria bind fibronectin through a specific receptor and it has been suggested that this binding may serve as a mechanism of tissue adherence [2]. The primary binding site for Staphylococci [3],

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Cloning and expression of the gene for a fibronectin-binding protein from Staphylococcus aureus.

Cited by 235 publications

References 24 publications

Dynamics of the interaction between a fibronectin molecule and a living bacterium under mechanical force

Dynamics of the interaction between a fibronectin molecule and a living bacterium under mechanical force

Nucleotide sequence of the gene for a fibronectin-binding protein from Staphylococcus aureus: use of this peptide sequence in the synthesis of biologically active peptides.

Multiple binding sites in fibronectin and the staphylococcal fibronectin receptor

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