Gram-positive pathogens such as staphylococci contain multiple cell wall-anchored proteins that serve as an interface between the microbe and its environment. Some of these proteins act as adhesins and mediate bacterial attachment to host tissues. SdrG is a cell wall-anchored adhesin from Staphylococcus epidermidis that binds to the Bbeta chain of human fibrinogen (Fg) and is necessary and sufficient for bacterial attachment to Fg-coated biomaterials. Here, we present the crystal structures of the ligand binding region of SdrG as an apoprotein and in complex with a synthetic peptide analogous to its binding site in Fg. Analysis of the crystal structures, along with mutational studies of both the protein and of the peptide, reveals that SdrG binds to its ligand with a dynamic "dock, lock, and latch" mechanism. We propose that this mechanism represents a general mode of ligand binding for structurally related cell wall-anchored proteins of gram-positive bacteria.

The shoot apical meristem (SAM) of seed plants is the site at which lateral organs are formed. Once organ primordia initiate from the SAM, they establish polarity along the adaxial-abaxial, proximodistal and mediolateral axes. Among these three axes, the adaxial-abaxial polarity is of primary importance in leaf patterning. In leaf development, once the adaxial-abaxial axis is established within leaf primordia, it provides cues for proper lamina growth and asymmetric development. It was reported previously that the Arabidopsis ASYMMETRIC LEAVES1 (AS1) and ASYMMETRIC LEAVES2 (AS2) genes are two key regulators of leaf polarity. In this work, we demonstrate a new function of the AS1 and AS2genes in the establishment of adaxial-abaxial polarity by analyzing as1 and as2 alleles in the Landsberg erecta(Ler) genetic background. We provide genetic evidence that the Arabidopsis ERECTA (ER) gene is involved in the AS1-AS2 pathway to promote leaf adaxial fate. In addition, we show that AS1 and AS2 bind to each other, suggesting that AS1 and AS2 may form a complex that regulates the establishment of leaf polarity. We also report the effects on leaf polarity of overexpression of the AS1 or AS2genes under the control of the cauliflower mosaic virus (CAMV) 35S promoter. Although plants with as1 and as2 mutations have very similar phenotypes, 35S::AS1/Ler and 35S::AS2/Lertransgenic plants showed dramatically different morphologies. A possible model of the AS1, AS2 and ER action in leaf polarity formation is discussed.

The collagens are a family of animal proteins containing segments of repeated Gly-Xaa-Yaa (GXY) motifs that form a characteristic triple-helical structure. Genes encoding proteins with repeated GXY motifs have also been reported in bacteria and phages; however, it is unclear whether these prokaryotic proteins can form a collagen-like triple-helical structure. Here we used two recently identified streptococcal proteins, Scl1 and Scl2, containing extended GXY sequence repeats as model proteins. First we observed that prior to heat denaturation recombinant Scl proteins migrated as homotrimers in gel electrophoresis with and without SDS. We next showed that the collagen-like domain of Scl is resistant to proteolysis by trypsin. We further showed that circular dichroism spectra of the Scl proteins contained features characteristic of collagen triple helices, including a positive maximum of ellipticity at 220 nm. FurthermorethetriplehelicesofScl1andScl2showedatemperature-dependent unfolding with melting temperatures of 36.4 and 37.6°C, respectively, which resembles those seen for collagens. We finally demonstrated by electron microscopy that the Scl proteins are organized into "lollipoplike" structures, similar to those seen in human proteins with collagenous domains. This implies that the repeated GXY tripeptide motif is a structural indicator of collagenlike triple helices in proteins from such phylogenetically distant sources as bacteria and humans.The collagens comprise a large family of structurally related proteins that are essential building elements of connective tissues in animals (1). In addition to contributing to the structural integrity of the tissue, collagens may also affect cell behavior, e.g. by interacting with specific cellular receptors. All collagens contain a triple-helical domain composed of three left-handed polyproline II-type chains wound around the central axis to form a right-handed superhelix (2). The segments of the collagen polypeptide forming a triple helix contain the repeating amino acid sequence Gly-Xaa-Yaa (GXY) in which a glycine residue occupies every third position, since only glycine is small enough to be accommodated at the center of the triple helix. The three chains are linked by hydrogen bonds between the backbone group, -NH (Gly) and the backbone carbonyl group (X position) of another chain, and by water-mediated interactions. The X and Y positions are frequently occupied by the imino acids proline and hydroxyproline, respectively. Hydroxyprolines are formed by post-translational hydroxylation of proline residues and are important in stabilizing the collagen triple helix. In humans, mutations that affect collagen triple helix formation and fiber assembly have serious pathological consequences often leading to death (3).The vertebrate collagens are classified into 19 types that are grouped in two major categories known as fibrillar (types I-III, V, and XI) and nonfibrillar (IV, VI-X, and XII-XIX) collagens. In addition, a variety of other mammalian proteins have been identif...

The structural basis for the association of eukaryotic and prokaryotic protein receptors and their triple-helical collagen ligand remains poorly understood. Here, we present the crystal structures of a high affinity subsegment of the Staphylococcus aureus collagen-binding CNA as an apoprotein and in complex with a synthetic collagen-like triple helical peptide. The apo-protein structure is composed of two subdomains (N1 and N2), each adopting a variant IgG-fold, and a long linker that connects N1 and N2. The structure is stabilized by hydrophobic interdomain interactions and by the N2 C-terminal extension that complements a b-sheet on N1. In the ligand complex, the collagen-like peptide penetrates through a spherical hole formed by the two subdomains and the N1-N2 linker. Based on these two structures we propose a dynamic, multistep binding model, called the 'Collagen Hug' that is uniquely designed to allow multidomain collagen binding proteins to bind their extended rope-like ligand.