RESUMEN
Streptococcus equi is the causative agent of the highly contagious disease "strangles" in equines and zoonotic meningitis in human. Spreading of infection in host tissues is thought to be facilitated by the bacterial gene encoded extracellular hyaluronate lyase (HL), which degrades hyaluronan (HA), chondroitin 6-sulfate, and dermatan sulfate of the extracellular matrix). The clinical strain S. equi 4047 however, lacks a functional extracellular HL. The prophages of S. equi and other streptococci encode intracellular HLs which are reported to partially degrade HA and do not cleave any other glycosaminoglycans. The phage HLs are thus thought to play a role limited to the penetration of streptococcal HA capsules, facilitating bacterial lysogenization and not in the bacterial pathogenesis. Here we systematically looked into the structure-function relationship of S. equi 4047 phage HL. Although HA is the preferred substrate, this HL has weak activity toward chondroitin 6-sulfate and dermatan sulfate and can completely degrade all of them. Even though the catalytic triple-stranded ß-helix domain of phage HL is functionally independent, its catalytic efficiency and specificity is influenced by the N-terminal domain. The phage HL also interacts with human transmembrane glycoprotein CD44. The above results suggest that the streptococci can use phage HLs to degrade glycosaminoglycans of the extracellular matrix for spreading virulence factors and toxins while utilizing the disaccharides as a nutrient source for proliferation at the site of infection.
Asunto(s)
Polisacárido Liasas/metabolismo , Profagos/enzimología , Streptococcus equi/virología , Proteínas Virales/metabolismo , Secuencias de Aminoácidos/genética , Animales , Dominio Catalítico , Sulfatos de Condroitina/metabolismo , Dermatán Sulfato/metabolismo , Electroforesis en Gel de Poliacrilamida , Polarización de Fluorescencia , Glicosaminoglicanos/metabolismo , Células HCT116 , Caballos , Humanos , Receptores de Hialuranos/metabolismo , Ácido Hialurónico/metabolismo , Cinética , Microscopía Confocal , Modelos Moleculares , Mutación , Polisacárido Liasas/química , Polisacárido Liasas/genética , Profagos/genética , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Especificidad por Sustrato , Proteínas Virales/química , Proteínas Virales/genéticaRESUMEN
Bacteriophages deploy lysins that degrade the bacterial cell wall and facilitate virus egress from the host. When applied exogenously, these enzymes destroy susceptible microbes and, accordingly, have potential as therapeutic agents. The most potent lysin identified to date is PlyC, an enzyme assembled from two components (PlyCA and PlyCB) that is specific for streptococcal species. Here the structure of the PlyC holoenzyme reveals that a single PlyCA moiety is tethered to a ring-shaped assembly of eight PlyCB molecules. Structure-guided mutagenesis reveals that the bacterial cell wall binding is achieved through a cleft on PlyCB. Unexpectedly, our structural data reveal that PlyCA contains a glycoside hydrolase domain in addition to the previously recognized cysteine, histidine-dependent amidohydrolases/peptidases catalytic domain. The presence of eight cell wall-binding domains together with two catalytic domains may explain the extraordinary potency of the PlyC holoenyzme toward target bacteria.
Asunto(s)
Enzimas/química , Fagos de Streptococcus/enzimología , Streptococcus equi/virología , Proteínas Virales/química , Cristalografía por Rayos X , Estructura Cuaternaria de Proteína , Estructura Terciaria de ProteínaRESUMEN
The continued evolution of bacterial pathogens has major implications for both human and animal disease, but the exchange of genetic material between host-restricted pathogens is rarely considered. Streptococcus equi subspecies equi (S. equi) is a host-restricted pathogen of horses that has evolved from the zoonotic pathogen Streptococcus equi subspecies zooepidemicus (S. zooepidemicus). These pathogens share approximately 80% genome sequence identity with the important human pathogen Streptococcus pyogenes. We sequenced and compared the genomes of S. equi 4047 and S. zooepidemicus H70 and screened S. equi and S. zooepidemicus strains from around the world to uncover evidence of the genetic events that have shaped the evolution of the S. equi genome and led to its emergence as a host-restricted pathogen. Our analysis provides evidence of functional loss due to mutation and deletion, coupled with pathogenic specialization through the acquisition of bacteriophage encoding a phospholipase A(2) toxin, and four superantigens, and an integrative conjugative element carrying a novel iron acquisition system with similarity to the high pathogenicity island of Yersinia pestis. We also highlight that S. equi, S. zooepidemicus, and S. pyogenes share a common phage pool that enhances cross-species pathogen evolution. We conclude that the complex interplay of functional loss, pathogenic specialization, and genetic exchange between S. equi, S. zooepidemicus, and S. pyogenes continues to influence the evolution of these important streptococci.
Asunto(s)
Evolución Molecular , Genes Bacterianos , Streptococcus equi/genética , Streptococcus equi/patogenicidad , Animales , Bacteriófagos/genética , Genoma , Caballos , Humanos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Streptococcus equi/virología , Streptococcus pyogenes/genética , VirulenciaRESUMEN
Streptococcus equi causes equine 'strangles'. Hyaluronate lyases, which degrade connective tissue hyaluronan and chondroitins, are thought to facilitate streptococcal invasion of the host. However, prophage-encoded hyaluronate lyases are hyaluronan-specific and are thought to be primarily involved in the degradation of the hyaluronan capsule of streptococci during bacteriophage infection. To understand the role of prophage-encoded hyaluronate lyases further, we have biochemically characterized such a hyaluronate lyase, SEQ2045 from S. equi, and have shown that it is produced during equine infection. Prophage-encoded hyaluronan-specific hyaluronate lyases may therefore play a more direct role in disease pathogenesis than previously thought.
Asunto(s)
Enfermedades de los Caballos/microbiología , Ácido Hialurónico/metabolismo , Polisacárido Liasas/metabolismo , Profagos/enzimología , Infecciones Estreptocócicas/veterinaria , Fagos de Streptococcus/enzimología , Streptococcus equi/virología , Secuencia de Aminoácidos , Animales , Caballos , Datos de Secuencia Molecular , Polisacárido Liasas/química , Polisacárido Liasas/genética , Alineación de Secuencia , Infecciones Estreptocócicas/microbiología , Streptococcus equi/química , Streptococcus equi/enzimología , Streptococcus equi/genética , Proteínas Virales/química , Proteínas Virales/genética , Proteínas Virales/metabolismoRESUMEN
Streptococcus equi (S. equi subsp. equi) is widely believed to have evolved from an ancestral strain of S. zooepidemicus (S. equi subsp. zooepidemicus) based on high sequence homology. A striking difference is the absence of phage sequences from S. zooepidemicus. In this study we show that the receptor for SeP9, a temperate bacteriophage of S. equi, is the Lancefield group C carbohydrate. However, although SeP9 binds to group C carbohydrate from S. zooepidemicus, it appears not to replicate and produce plaques.