Functional Mapping of an Oligomeric Autotransporter Adhesin of Aggregatibacter actinomycetemcomitans

Abstract

Extracellular matrix protein adhesin A (EmaA) is a 202-kDa nonfimbrial adhesin, which mediates the adhesion of the oral pathogen Aggregatibacter actinomycetemcomitans to collagen. EmaA oligomers form surface antenna-like protrusions consisting of a long helical rod with an ellipsoidal ending. The functional analysis of in-frame emaA deletion mutants has located the collagen binding activity to the amino terminus of the protein corresponding to amino acids 70 to 386. The level of collagen binding of this deletion mutant was comparable to the emaA mutant strain. Transmission electron microscopy studies indicate that the first 330 amino acids of the mature protein form the ellipsoidal ending of the EmaA protrusions, where the activity resides. Amino acid substitution analysis within this sequence has identified a critical amino acid, which is essential for the formation of the ellipsoidal ending and for collagen binding activity.The interaction with extracellular matrix (ECM) proteins of the host is an established virulence determinant in many bacterial pathogens (13). Bacteria express several surface proteinaceous structures for adhesion to ECM proteins. These structures are categorized as fimbrial and nonfimbrial adhesins and can be visualized in negatively stained images of bacteria by transmission electron microscopy (TEM) (12,32,40). Fimbrial adhesins are typically long structures with lengths of 0.3 to 1 m (5) composed of multiple copies of homo-or heteropolymeric subunits (14). Nonfimbrial adhesins form smaller structures, up to 0.3 m in length, which consist of a single protein monomer, or small homo-oligomers anchored to the surface of the bacteria (12,13,29).Homo-oligomeric adhesins compose a subclass of the type V autotransporter secretion system, which include types V a , V b ,

Glycosylation of the Collagen Adhesin EmaA ofAggregatibacter actinomycetemcomitansIs Dependent upon the Lipopolysaccharide Biosynthetic Pathway

“…The EmaA of serotype b A. actinomycetemcomitans is a 202-kDa protein that forms the antennae-like appendages found on the surface of A. actinomycetemcomitans and is required for collagen binding (40). The appendages are composed of three EmaA monomers that oligomerize to form an ellipsoidal structure required for the collagen binding activity (56,57). The ellipsoidal structure corresponds to the amino termini of the proteins and is located at the distal end of a long stalk domain that is attached to the outer membrane by the carboxyl termini (57).…”

“…The appendages are composed of three EmaA monomers that oligomerize to form an ellipsoidal structure required for the collagen binding activity (56,57). The ellipsoidal structure corresponds to the amino termini of the proteins and is located at the distal end of a long stalk domain that is attached to the outer membrane by the carboxyl termini (57). The carboxyl termini of the proteins assume ␤-barrel structures required for pore formation and translocation of the molecules through the outer membrane, similar to those of other type V c autotransporter proteins (14).…”

Ultrastructural Analysis of the Rugose Cell Envelope of a Member of the Pasteurellaceae Family

“…1D). Images of the wild-type strain show grooves on the bacterial surface and also the presence of the collagen binding adhesin, extracellular matrix adhesin A (EmaA), that forms antenna-like structures protruding from the bacterial surface (13,22,30,31). Our earlier studies revealed that convolutions in A. actinomycetemcomitans are associated with a protein called morphogenesis protein C (MorC), which appears to be located in the inner membrane (29).…”

“…However, the loss of the rugosity does not seem to influence the presentation of all virulence determinants on the outer bacterial surface. EmaA (extracellular matrix adhesin A), a trimeric autotransporter collagen binding adhesin that forms antenna-like appendages on the surface of the bacterium (13,22,30,31), is present in both wild-type and morC mutant strains, thus indicating that not all outer membrane transport systems are affected. All of these observations raise fundamental questions about the specific role of the outer membrane morphology and the spatial relationship between outer and inner membranes in bacteria with a native rugose phenotype.…”

Stick to Your Gums

“…EmaA is structurally distinct from the other two auto-transporters, with coiled-coil, glycosylated monomers trimerizing to form antenna-like protrusions on the bacterial cell surface. These comprise a stalk of approximately 150 nm that terminates in an ellipsoidal cap, within which the collagenbinding domain is located (Ruiz et al, 2006;Yu et al, 2008;Tang and Mintz, 2010). Specific amino acid residues within the stalk have been shown to introduce bends within the structure, conferring flexibility to ensure optimal adhesion to collagen (Yu et al, 2009).…”