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Streptococcus mutans antigen I/II (AgI/II) is a cell surface-localized protein adhesin that interacts with salivary components within the salivary pellicle. AgI/II contributes to virulence and has been studied as an immunological and structural target, but a fundamental understanding of its underlying architecture has been lacking. Here we report a high-resolution (1.8 Å) crystal structure of the A 3 VP 1 fragment of S. mutans AgI/II that demonstrates a unique fibrillar form (155 Å) through the interaction of two noncontiguous regions in the primary sequence. The A 3 repeat of the alanine-rich domain adopts an extended α-helix that intertwines with the P 1 repeat polyproline type II (PPII) helix to form a highly extended stalk-like structure heretofore unseen in prokaryotic or eukaryotic protein structures. Velocity sedimentation studies indicate that fulllength AgI/II that contains three A/P repeats extends over 50 nanometers in length. Isothermal titration calorimetry revealed that the high-affinity association between the A 3 and P 1 helices is enthalpically driven. Two distinct binding sites on AgI/II to the host receptor salivary agglutinin (SAG) were identified by surface plasmon resonance (SPR). The current crystal structure reveals that AgI/II family proteins are extended fibrillar structures with the number of alanine-and proline-rich repeats determining their length.bacterial adhesion | dental caries | Streptococcus | x-ray crystallography | fibrous proteins S treptococcus mutans is the causative agent of human dental caries (1) and its protein adhesin antigen I/II (AgI/II) is a known target of protective immunity (2). AgI/II family molecules are expressed by numerous oral streptococci (3) and homologs have also been identified in the invasive pathogens Streptococcus pyogenes and Streptococcus agalactiae (4) (Fig. S1). In addition to mediating adhesion to the tooth surface (5), AgI/II influences biofilm formation (6), promotes collagen-dependent bacterial invasion of dentin (7), and mediates adherence to human epithelial cells (8). Elimination of AgI/II results in decreased virulence (9), but despite three decades of study, a mechanistic understanding of the functional properties of the molecule has been stymied by a lack of understanding of its structure.Originally identified as AgI/II (10) (also called P1, PAc, or SpaP), members of this protein family contain between 1310 and 1653 amino acids (aa) beginning with an amino-terminal signal motif that directs secretion, followed by the A, V, and P regions (Fig. 1A). The A region typically consists of 3-4 alanine-rich repeats (82 residues each) with 23-30% alanine content. The P region has 3-4 proline-rich repeats (39 residues each) with ∼35% proline content. Nested between the A and P repeats is a segment commonly referred to as the V or variable region, which contains within it a stretch of ∼100 amino acids where most of the sequence variation among S. mutans AgI/II molecules is clustered (11). The crystal structure of the V region adopts a globular β-s...

The Streptococcus mutans antigen I/II (AgI/II) is a cell surface-localized protein that adheres to salivary components and extracellular matrix molecules. Here we report the 2.5 Å resolution crystal structure of the complete C-terminal region of AgI/ II. The C-terminal region is comprised of three major domains: C 1 , C 2 , and C 3 . Each domain adopts a DE-variant IgG fold, with two ␤-sheets whose A and F strands are linked through an intramolecular isopeptide bond. The adherence of the C-terminal AgI/II fragments to the putative tooth surface receptor salivary agglutinin (SAG), as monitored by surface plasmon resonance, indicated that the minimal region of binding was contained within the first and second DE-variant-IgG domains (C 1 and C 2 ) of the C terminus. The minimal C-terminal region that could inhibit S. mutans adherence to SAG was also confirmed to be within the C 1 and C 2 domains. Competition experiments demonstrated that the C-and N-terminal regions of AgI/II adhere to distinct sites on SAG. A cleft formed at the intersection between these C 1 and C 2 domains bound glucose molecules from the cryo-protectant solution, revealing a putative binding site for its highly glycosylated receptor SAG. Finally, electron microscopy images confirmed the elongated structure of AgI/II and enabled building a composite tertiary model that encompasses its two distinct binding regions.Dental caries (also called tooth decay or dental cavities) is a ubiquitous worldwide disease that affects humans of all age groups. Streptococcus mutans, a primary etiological agent of human dental caries (1) and an increasingly recognized cause of bacterial endocarditis (2), adheres to proteins contained within the salivary pellicle on the tooth surface, the extracellular matrix, and other microbial species (3). Antigen I/II (AgI/II, 2 also known as P1, B, SpaP, or PAc) of S. mutans has been implicated in bacterial adherence to constituents of the salivary pellicle (4, 5) and has been studied for the past three decades as a target for protective immunity against dental caries. Apart from adherence, AgI/II influences biofilm formation (6) and promotes platelet aggregation (7), collagen-dependent bacterial invasion of dentin (8), and cariogenicity (9). Although AgI/II was initially discovered on oral streptococci, it has also been identified in members of the Group A and Group B streptococci (10), suggesting a role for this adhesin in a variety of species.The AgI/II family proteins range from 140 to 180 kDa in predicted size and have a primary sequence composed of multiple conserved regions (Fig. 1b). Toward the N terminus, repeated sequences of high alanine content constitute the alanine-rich region followed by a segment commonly referred to as the variable (V) region. Further C-terminal in the sequence is a region of high proline content that forms a repetitive prolinerich region. Following the AgI/II proline-rich region is a C-terminal region (60 kDa or 550 amino acids), which is the most conserved region of AgI/II, with 62% identity ...

Acquired resistance to androgen receptor (AR)-targeted therapies compels the development of novel treatment strategies for castration-resistant prostate cancer (CRPC). Here, we report a profound effect of endostatin on prostate cancer cells by efficient intracellular trafficking, direct interaction with AR, reduction of nuclear AR level, and down-regulation of AR-target gene transcription. Structural modeling followed by functional analyses further revealed that phenylalanine-rich α1-helix in endostatin-which shares structural similarity with noncanonical nuclear receptor box in ARantagonizes AR transcriptional activity by occupying the activation function (AF)-2 binding interface for coactivators and N-terminal AR AF-1. Together, our data suggest that endostatin can be recognized as an endogenous AR inhibitor that impairs receptor function through protein-protein interaction. These findings provide new insights into endostatin whose antitumor effect is not limited to inhibiting angiogenesis, but can be translated to suppressing AR-mediated disease progression in CRPC.endostatin | androgen receptor | prostate cancer | chemoresistance

The high-resolution structure of glucan binding protein C (GbpC) at 1.14 Å, a sucrose-dependent virulence factor of the dental caries pathogen , has been determined. GbpC shares not only structural similarities with the V regions of AgI/II and SspB but also functional adherence to salivary agglutinin (SAG) and its scavenger receptor cysteine-rich domains (SRCRs). This is not only a newly identified function for GbpC but also an additional fail-safe binding mechanism for Despite the structural similarities with antigen I/II (AgI/II) and SspB of, GbpC remains unique among these surface proteins in its propensity to adhere to dextran/glucans. The complex crystal structure of GbpC with dextrose (β-d-glucose; Protein Data Bank ligand BGC) highlights exclusive structural features that facilitate this interaction with dextran. Targeted deletion mutant studies on GbpC's divergent loop region in the vicinity of a highly conserved calcium binding site confirm its role in biofilm formation. Finally, we present a model for adherence to dextran. The structure of GbpC highlights how artfully microbes have engineered the lectin-like folds to broaden their functional adherence repertoire.