Structure of the K141 capsular polysaccharide produced by Acinetobacter baumannii isolate KZ1106 that carries KL141 at the chromosomal K locus.

The structure of the K141 type capsular polysaccharide (CPS) produced by Acinetobacter baumannii KZ1106, a clinical isolate recovered from Kazakhstan in 2016, was established by sugar analyses and one- and two-dimensional 1H and 13C NMR spectroscopy. The CPS was shown to consist of branched tetrasaccharide repeating units (K-units) with the following structure: This structure was found to be consistent with the genetic content of the KL141 CPS biosynthesis gene cluster at the chromosomal K locus in the KZ1106 whole genome sequence. Assignment of the encoded enzymes allowed the first sugar of the K unit to be identified, which revealed that the ß-d-GlcpNAc-(1→3)-d-GlcpNAc bond is the linkage between K-units formed by the WzyKL141 polymerase.

The Acinetobacter baumannii K239 capsular polysaccharide includes heptasaccharide units that are structurally related to K86 but joined by different linkages formed by different Wzy polymerases.

The K239 type capsular polysaccharide (CPS) isolated from Acinetobacter baumannii isolate MAR19-4435 was studied by sugar analysis, one- and two-dimensional 1H and 13C NMR spectroscopy. K239 consists of branched heptasaccharide repeats (K-units) comprised of five residues of l-rhamnose (l-Rhap), and one residue each of d-glucuronic acid (d-GlcpA) and N-acetyl-d-glucosamine (d-GlcpNAc). The structure of K239 is closely related to that of the A. baumannii K86 CPS type, though the two differ in the 2,3-substitution patterns on the l-Rhap residue that is involved in the linkage between K-units in the CPS polymer. This structural difference was attributed to the presence of a gtr221 glycosyltransferase gene and a wzyKL239 polymerase gene in KL239 that replaces the gtr80 and wzyKL86 genes in the KL86 CPS biosynthesis gene cluster. Comparison of the two structures established the role of a novel WzyKL239 polymerase encoded by KL239 that forms the ß-d-GlcpNAc-(1→2)-l-Rhap linkage between K239 units. A. baumannii MAR19-4435 was found to be non-susceptible to infection by the APK86 bacteriophage, which encodes a depolymerase that specifically cleaves the linkage between K-units in the K86 CPS, indicating that the difference in 2,3-substitution of l-Rhap influences the susceptibility of this isolate to bacteriophage activity.

New Obolenskvirus Phages Brutus and Scipio: Biology, Evolution, and Phage-Host Interaction.

Two novel virulent phages of the genus Obolenskvirus infecting Acinetobacter baumannii, a significant nosocomial pathogen, have been isolated and studied. Phages Brutus and Scipio were able to infect A. baumannii strains belonging to the K116 and K82 capsular types, respectively. The biological properties and genomic organization of the phages were characterized. Comparative genomic, phylogenetic, and pangenomic analyses were performed to investigate the relationship of Brutus and Scipio to other bacterial viruses and to trace the possible origin and evolutionary history of these phages and other representatives of the genus Obolenskvirus. The investigation of enzymatic activity of the tailspike depolymerase encoded in the genome of phage Scipio, the first reported virus infecting A. baumannii of the K82 capsular type, was performed. The study of new representatives of the genus Obolenskvirus and mechanisms of action of depolymerases encoded in their genomes expands knowledge about the diversity of viruses within this taxonomic group and strategies of Obolenskvirus-host bacteria interaction.

Revised structure of the polysaccharide from Acinetobacter baumannii LUH5551 assigned as the K63 type capsular polysaccharide.

K63 capsular polysaccharide produced by Acinetobacter baumannii isolate LUH5551 (previously designated isolate O24) was re-examined using sugar analysis, Smith degradation, and one- and two-dimensional 1H and 13C NMR spectroscopy. Though previously reported as O24 consisting of linear tetrasaccharide units that include a 7-acetamido-5-acylamino form of 8-epilegionaminic acid [8eLeg5R7Ac, acylated at C5 with (S)-3-hydroxybutanoyl or acetyl (1:1)], the elucidated structure of the K63 type capsule was found to include a derivative of 5,7-diamino-3,5,7,9-tetradeoxy-d-glycero-d-galacto-non-2-ulosonic (legionaminic) acid, Leg5Ac7R, where R is either (S)-3-hydroxybutanoyl or an acetyl group (∼1:1 ratio). This finding is consistent with the presence of the lgaABCHIFG gene module for Leg5Ac7R biosynthesis in the KL63 gene cluster at the capsular polysaccharide (CPS) biosynthesis K locus in the LUH5551 genome. The glycosyltransferases (Gtrs) and Wzy polymerase encoded by KL63 were assigned to linkages in the linear K63 tetrasaccharide unit and linkage of the K63 units.

Depolymerisation of the Klebsiella pneumoniae Capsular Polysaccharide K21 by Klebsiella Phage K5.

Klebsiella pneumoniae is a pathogen associated with various infection types, which often exhibits multiple antibiotic resistance. Phages, or bacterial viruses, have an ability to specifically target and destroy K. pneumoniae, offering a potential means of combatting multidrug-resistant infections. Phage enzymes are another promising therapeutic agent that can break down bacterial capsular polysaccharide, which shields K. pneumoniae from the immune response and external factors. In this study, Klebsiella phage K5 was isolated; this phage is active against Klebsiella pneumoniae with the capsular type K21. It was demonstrated that the phage can effectively lyse the host culture. The adsorption apparatus of the phage has revealed two receptor-binding proteins (RBPs) with predicted polysaccharide depolymerising activity. A recombinant form of both RBPs was obtained and experiments showed that one of them depolymerised the capsular polysaccharide K21. The structure of this polysaccharide and its degradation fragments were analysed. The second receptor-binding protein showed no activity on capsular polysaccharide of any of the 31 capsule types tested, so the substrate for this enzyme remains to be determined in the future. Klebsiella phage K5 may be considered a useful agent against Klebsiella infections.

The Acinetobacter baumannii K70 and K9 capsular polysaccharides consist of related K-units linked by the same Wzy polymerase and cleaved by the same phage depolymerases.

IMPORTANCE: Bacteriophage show promise for the treatment of Acinetobacter baumannii infections that resist all therapeutically suitable antibiotics. Many tail-spike depolymerases encoded by phage that are able to degrade A. baumannii capsular polysaccharide (CPS) exhibit specificity for the linkage present between K-units that make up CPS polymers. This linkage is formed by a specific Wzy polymerase, and the ability to predict this linkage using sequence-based methods that identify the Wzy at the K locus could assist with the selection of phage for therapy. However, little is known about the specificity of Wzy polymerase enzymes. Here, we describe a Wzy polymerase that can accommodate two different but similar sugars as one of the residues it links and phage depolymerases that can cleave both types of bond that Wzy forms.

Friunavirus Phage-Encoded Depolymerases Specific to Different Capsular Types of Acinetobacter baumannii.

Acinetobacter baumannii is a critical priority nosocomial pathogen that produces a variety of capsular polysaccharides (CPSs), the primary receptors for specific depolymerase-carrying phages. In this study, the tailspike depolymerases (TSDs) encoded in genomes of six novel Friunaviruses, APK09, APK14, APK16, APK86, APK127v, APK128, and one previously described Friunavirus phage, APK37.1, were characterized. For all TSDs, the mechanism of specific cleavage of corresponding A. baumannii capsular polysaccharides (CPSs) was established. The structures of oligosaccharide fragments derived from K9, K14, K16, K37/K3-v1, K86, K127, and K128 CPSs degradation by the recombinant depolymerases have been determined. The crystal structures of three of the studied TSDs were obtained. A significant reduction in mortality of Galleria mellonella larvae infected with A. baumannii of K9 capsular type was shown in the example of recombinant TSD APK09_gp48. The data obtained will provide a better understanding of the interaction of phage-bacterial host systems and will contribute to the formation of principles of rational usage of lytic phages and phage-derived enzymes as antibacterial agents.

5,7-Diamino-3,5,7,9-tetradeoxynon-2-ulosonic Acids in the Capsular Polysaccharides of Acinetobacter baumannii.

The polysaccharide capsule surrounding bacterial cell plays an important role in pathogenesis of infections caused by the opportunistic pathogen Acinetobacter baumannii by providing protection from external factors. The structures of the capsular polysaccharide (CPS) produced by A. baumannii isolates and the corresponding CPS biosynthesis gene clusters are highly diverse, although many of them are related. Many types of A. baumannii CPSs contain isomers of 5,7-diamino-3,5,7,9-tetradeoxynon-2-ulosonic acid (DTNA). Three of these isomers, namely acinetaminic acid (l-glycero-l-altro isomer), 8-epiacinetaminic acid (d-glycero-l-altro isomer), and 8-epipseudaminic acid (d-glycero-l-manno isomer), have not been found so far in naturally occurring carbohydrates from other species. In A. baumannii CPSs, DTNAs carry N-acyl substituents at positions 5 and 7; in some CPSs, both N-acetyl and N-(3-hydroxybutanoyl) groups are present. Remarkably, pseudaminic acid carries the (R)-isomer and legionaminic acid carries the (S)-isomer of the 3-hydroxybutanoyl group. The review addresses the structure and genetics of biosynthesis of A. baumannii CPSs containing di-N-acyl derivatives of DTNA.

Structural and Serological Characterization of the O Antigen of Proteus mirabilis Clinical Isolates Classified into a New Proteus Serogroup, O84.

Two closely related Proteus mirabilis smooth strains, Kr1 and Ks20, were isolated from wound and skin samples, respectively, of two infected patients in central Poland. Serological tests, using the rabbit Kr1-specific antiserum, revealed that both strains presented the same O serotype. Their O antigens are unique among the Proteus O serotypes, which had been described earlier, as they were not recognized in an enzyme-linked immunosorbent assay (ELISA) by a set of Proteus O1-O83 antisera. Additionally, the Kr1 antiserum did not react with O1-O83 lipopolysaccharides (LPSs). The O-specific polysaccharide (OPS, O antigen) of P. mirabilis Kr1 was obtained via the mild acid degradation of the LPSs, and its structure was established via a chemical analysis and one- and two-dimensional 1H and 13C nuclear magnetic resonance (NMR) spectroscopy applied to both initial and O-deacetylated polysaccharides, where most ß-2-acetamido-2-deoxyglucose (N-acetylglucosamine) (GlcNAc) residues are non-stoichiometrically O-acetylated at positions 3, 4, and 6 or 3 and 6, and a minority of α-GlcNAc residues are 6-O-acetylated. Based on the serological features and chemical data, P. mirabilis Kr1 and Ks20 were proposed as candidates to a new successive O-serogroup in the genus Proteus, O84, which is another example of new Proteus O serotypes identified lately among serologically differentiated Proteus bacilli infecting patients in central Poland.

Loss of a Branch Sugar in the Acinetobacter baumannii K3-Type Capsular Polysaccharide Due To Frameshifts in the gtr6 Glycosyltransferase Gene Leads To Susceptibility To Phage APK37.1.

The type of capsular polysaccharide (CPS) on the cell surface of Acinetobacter baumannii can determine the specificity of lytic bacteriophage under consideration for therapeutic use. Here, we report the isolation of a phage on an extensively antibiotic resistant ST2 A. baumannii isolate AB5001 that carries the KL3 CPS biosynthesis gene cluster predicting a K3-type CPS. As the phage did not infect isolates carrying KL3 or KL22 and known to produce K3 CPS, the structure of the CPS isolated from A. baumannii AB5001 was determined. AB5001 produced a variant CPS form, K3-v1, that lacks the ß-d-GlсpNAc side chain attached to the d-Galp residue in the K3 structure. Inspection of the KL3 sequence in the genomes of AB5001 and other phage-susceptible isolates with a KL3 locus revealed single-base deletions in gtr6, causing loss of the Gtr6 glycosyltransferase that adds the missing d-GlсpNAc side chain to the K3 CPS. Hence, the presence of this sugar profoundly restricts the ability of the phage to digest the CPS. The 41-kb linear double-stranded DNA (dsDNA) phage genome was identical to the genome of a phage isolated on a K37-producing isolate and thus was named APK37.1. APK37.1 also infected isolates carrying KL116. Consistent with this, K3-v1 resembles the K37 and K116 structures. APK37.1 is a Friunavirus belonging to the Autographiviridae family. The phage-encoded tail spike depolymerase DpoAPK37.1 was not closely related to Dpo encoded by other sequenced Friunaviruses, including APK37 and APK116. IMPORTANCE Lytic bacteriophage have potential for the treatment of otherwise untreatable extensively antibiotic-resistant bacteria. For Acinetobacter baumannii, most phage exhibit specificity for the type of capsular polysaccharide (CPS) produced on the cell surface. However, resistance can arise via mutations in CPS genes that abolish this phage receptor. Here, we show that single-base deletions in a CPS gene result in alteration of the final structure rather than deletion of the capsule layer and hence affect the ability of a newly reported podophage to infect strains producing the K3 CPS.

Cataloging natural sialic acids and other nonulosonic acids (NulOs), and their representation using the Symbol Nomenclature for Glycans.

Nonulosonic acids or non-2-ulosonic acids (NulOs) are an ancient family of 2-ketoaldonic acids (α-ketoaldonic acids) with a 9-carbon backbone. In nature, these monosaccharides occur either in a 3-deoxy form (referred to as "sialic acids") or in a 3,9-dideoxy "sialic-acid-like" form. The former sialic acids are most common in the deuterostome lineage, including vertebrates, and mimicked by some of their pathogens. The latter sialic-acid-like molecules are found in bacteria and archaea. NulOs are often prominently positioned at the outermost tips of cell surface glycans, and have many key roles in evolution, biology and disease. The diversity of stereochemistry and structural modifications among the NulOs contributes to more than 90 sialic acid forms and 50 sialic-acid-like variants described thus far in nature. This paper reports the curation of these diverse naturally occurring NulOs at the NCBI sialic acid page (https://www.ncbi.nlm.nih.gov/glycans/sialic.html) as part of the NCBI-Glycans initiative. This includes external links to relevant Carbohydrate Structure Databases. As the amino and hydroxyl groups of these monosaccharides are extensively derivatized by various substituents in nature, the Symbol Nomenclature For Glycans (SNFG) rules have been expanded to represent this natural diversity. These developments help illustrate the natural diversity of sialic acids and related NulOs, and enable their systematic representation in publications and online resources.

Protective Capacity of Monoclonal Antibodies against Acinetobacter baumannii K9 Capsular Polysaccharide.

Acinetobacter baumannii is an antibiotic-resistant opportunistic pathogen, one of the main causes of hospital infections. There is an urgent need for the development of therapy strategies which are not based on antibiotics. Hybridoma technology was used to obtain monoclonal antibodies. The antibodies were characterized by enzyme immunoassay and fluorescence microscopy according to their ability to opsonize A. baumannii and to protect model animals from infection upon intraperitoneal and pulmonary injection. Monoclonal antibodies (MAbs), IgG, against the K9 capsular polysaccharide (CPS) of A. baumannii were prepared using a glycoconjugate, synthesized by squaric-acid chemistry, consisting of two CPS K9 monomer units and a carrier protein. The MAbs were highly specific, stained the bacterial surface, allowed detection of A. baumannii in infected lung tissue, effectively opsonized the bacteria at nanogram concentrations (up to 1.5 ng/mL for CPS-407), and demonstrated a high ability to protect an organism against bacterial infection upon intraperitoneal and lung injection. In intraperitoneal infection of a mouse model with A. baumannii K9, the CPS-407 antibody protected at a dose of 25 µg/mouse. When bacteria were injected into the lung, MAb therapy prevented infection of the body and led to a significant reduction of the bacterial load in infected tissues. IMPORTANCE MAbs detected A. baumannii in infected lung tissue, effectively opsonized bacteria, and protected model animals from infection.

Complete chemical structure of the K135 capsular polysaccharide produced by Acinetobacter baumannii RES-546 that contains 5,7-di-N-acetyl-8-epipseudaminic acid.

A structurally diverse capsular polysaccharide (CPS) in the outer cell envelope plays an important role in the virulence of the important bacterial pathogen, Acinetobacter baumannii. More than 75 different CPS structures have been determined for the species to date, and many CPSs include isomers of a higher sugar, namely 5,7-diamino-3,5,7,9-tetradeoxynon-2-ulosonic acid. Recently, a novel isomer having the d-glycero-l-manno configuration (5,7-di-N-acetyl-8-epipseudaminic acid; 8ePse5Ac7Ac) has been identified in the CPS from A. baumannii clinical isolate RES-546 [Carbohydr. Res. 513 (2022) 108,531]. Here, the complete chemical structure of this CPS, designated K135, was elucidated. The CPS was found to have a branched tetrasaccharide K unit and to include the higher sugar as part of a 8ePse5Ac7Ac-(2 â†’ 6)-α-Gal disaccharide branching from a →3)-α-D-GlcpNAc-(1 â†’ 3)-ß-D-GlcpNAc-(1→ main chain. Assignment of glycosyltransferases encoded by the CPS biosynthesis gene cluster in the RES-546 genome enabled the first sugar of the K unit, and hence the topology of the K135 CPS, to be determined.

Comparison of the therapeutic potential of bacteriophage KpV74 and phage-derived depolymerase (ß-glucosidase) against Klebsiella pneumoniae capsular type K2.

Bacteriophages and phage polysaccharide-degrading enzymes (depolymerases) are garnering attention as possible alternatives to antibiotics. Here, we describe the antimicrobial properties of bacteriophage KpV74 and phage depolymerase Dep_kpv74 specific to the hypervirulent Klebsiella pneumoniae of the K2 capsular type. The depolymerase Dep_kpv74 was identified as a specific glucosidase that cleaved the K2 type capsular polysaccharides of the K. pneumoniae by a hydrolytic mechanism. This depolymerase was effective against thigh soft tissue K. pneumoniae infection in mice without inducing adverse behavioral effects or toxicity. The depolymerase efficiency was similar to or greater than the bacteriophage efficiency. The phage KpV74 had a therapeutic effect only for treating the infection caused by the phage-propagating K. pneumoniae strain and was completely inactive against the infection caused by the K. pneumoniae strain that did not support phage multiplication. The depolymerase was effective in both cases. A mutant resistant to phage and depolymerase was isolated during the treatment of mice with bacteriophage. A confirmed one-base deletion in the flippase-coding wzx gene of this mutant is assumed to affect the polysaccharide capsule, abolishing the KpV74 phage adsorption and reducing the K. pneumoniae virulence.

Pectobacterium versatile Bacteriophage Possum: A Complex Polysaccharide-Deacetylating Tail Fiber as a Tool for Host Recognition in Pectobacterial Schitoviridae.

Novel, closely related phages Possum and Horatius infect Pectobacterium versatile, a phytopathogen causing soft rot in potatoes and other essential plants. Their properties and genomic composition define them as N4-like bacteriophages of the genus Cbunavirus, a part of a recently formed family Schitoviridae. It is proposed that the adsorption apparatus of these phages consists of tail fibers connected to the virion through an adapter protein. Tail fibers possess an enzymatic domain. Phage Possum uses it to deacetylate O-polysaccharide on the surface of the host strain to provide viral attachment. Such an infection mechanism is supposed to be common for all Cbunavirus phages and this feature should be considered when designing cocktails for phage control of soft rot.

Immune Response to Conjugates of Fragments of the Type K9 Capsular Polysaccharide of Acinetobacter baumannii with Carrier Proteins.

The clonal bacterial species Acinetobacter baumannii is an emerging multidrug-resistant pathogen which causes high-lethality infections. Cells of A. baumannii are surrounded by the type-specific capsular polysaccharide (CPS), which provides resistance to the protective mechanisms of the host and is considered a target for immunization. The conjugates of three inert carrier proteins and A. baumannii type K9 CPS fragments, which contained various numbers of oligosaccharide repeats (K-units), were synthesized by periodate oxidation and squaric acid chemistry. The conjugates were applied to immunize mice, and chemical synthesis by squaric acid was shown to significantly improve the immunogenic properties of glycoconjugate. In BALB/c mice, IgG antibodies were predominant among type K9 CPS reactive antibodies, and their total content was several times higher than that of IgM. Immune sera were characterized by their opsonization ability during practically the entire lives of the experimental mice. The sera were cross-reactive, but the highest specificity was observed against the antigen (type K9 CPS) used for immunization. The immunization of BALB/c and ICR-1 mice with a glycoconjugate without adjuvants led to varying degrees of stimulation of IL-10, IL-17A, and TNF-α production, but not IL-4 production in the ICR-1 mice. This is in contrast to the BALB/c mice, in which γ-IFN production was also activated. The protective effectiveness of the glycoconjugates obtained by squaric acid chemistry was demonstrated by experiments that involved challenging immunized and nonimmunized animals with a lethal dose of A. baumannii K9. IMPORTANCE Immunization by glycoconjugates with A. baumannii type K9 CPS fragments induced a high level of antibodies (predominantly IgG) in sera, which reacted specifically with the CPS of A. baumannii type K9, as well as a long immunological memory. The sera of immunized animals efficiently opsonized A. baumannii type K9. Immunization resulted in the balanced production of pro/anti-inflammatory lymphokines and protective antibodies to ensure the survival of the mice infected with A. baumannii. The level of specific antibodies was sufficient to provide protective immunity against the challenge by A. baumannii, making this approach applicable in the development of vaccine preparations.

The K218 capsular polysaccharide produced by Acinetobacter baumannii isolate 52-249 includes 5,7-di-N-acetylpseudaminic acid linked by a KpsS3 glycosyltransferase.

Two acylated forms of the higher sugar, 5,7-diamino-3,5,7,9-tetradeoxy-l-glycero-l-manno-non-2-ulosonic acid called pseudaminic acid, Pse5Ac7Ac and Pse5Ac7RHb where R indicates (R)-3-hydroxybutanoyl, have been found to occur in many capsular polysaccharide (CPS) types produced by isolates of an important human pathogen, Acinetobacter baumannii. The presence of either a psaABCEDF or psaABCGHF gene module at the K locus (KL) for CPS biosynthesis determines the type of the variant produced. Here, an A. baumannii clinical isolate 52-249, recovered in 2015 in Moscow, Russia, was found to include a novel psaABCIJF gene module in the KL218 sequence at the K locus. The CPS from 52-249 was extracted and studied by sugar analysis and partial acid hydrolysis along with one- and two-dimensional 1H and 13C NMR spectroscopy. A branched tetrasaccharide repeating unit was identified, which included a →3)-α-d-Galp-(1→6)-α-d-GlcpNAc-(1→3)-ß-d-GalpNAc-(1→ main chain and Pse5Ac7Ac attached as a side branch, indicating that the psaABCIJF gene module is associated with synthesis of this variant. The K218 CPS was found to be structurally related to the K46 CPS of A. baumannii, and a comparison of the two structures enabled the assignment of glycosyltransferases. A KpsS3 protein for the α-(2→6) linkage of the Pse5Ac7Ac residue to D-Galp in K218 was identified.

Structure of the K98 capsular polysaccharide from Acinetobacter baumannii REV-1184 containing a cyclic pyruvic acid acetal.

The K98 capsular polysaccharide (CPS) from the Acinetobacter baumannii clinical isolate, REV-1184, was studied by sugar analysis and Smith degradation along with one- and two-dimensional 1H and 13C NMR spectroscopy and high-resolution electrospray ionization mass spectrometry. The CPS was found to consist of linear tetrasaccharide repeats (K-units) that include one residue each of d-GlcpNAc, d-GalpNAc, 2-acetamido-2-deoxy-d-galacturonic acid (d-GalpNAcA), and 2-acetamido-2,6-dideoxy-d-glucose (N-acetylquinovosamine, d-QuipNAc), with the GalpNAc residue decorated with a (R)-configurated 4,6-pyruvic acid acetal group. The CPS has a similar composition to that of A. baumannii K4 but the topology of the tetrasaccharide K-unit is different (linear in K98 versus branched in K4). This was due to a difference in sequence for the Wzy polymerases encoded by the CPS biosynthesis gene clusters KL98 and KL4, with the WzyK98 polymerase forming a ß-d-QuipNAc-(1→3)-d-GalpNAc linkage between the K98 units.

The K89 capsular polysaccharide produced by Acinetobacter baumannii LUH5552 consists of a pentameric repeat-unit that includes a 3-acetamido-3,6-dideoxy-d-galactose residue.

Acinetobacter baumannii isolate LUH5552 carries the KL89 capsule biosynthesis gene cluster. Capsular polysaccharide (CPS) isolated from LUH5552 was analyzed by sugar analysis, Smith degradation, and one- and two-dimensional 1H and 13C NMR spectroscopy. The K89 CPS structure has not been seen before in A. baumannii CPS structures resolved to date and includes a 3-acetamido-3,6-dideoxy-d-galactose (d-Fucp3NAc) residue which is rare amongst A. baumannii CPS. The K89 CPS has a →3)-α-d-GalpNAc-(1→3)-ß-d-GlcpNAc-(1→ main chain with a ß-d-Glcp-(1→2)-ß-d-Fucp3NAc-(1→6)-d-Glcp side branch that is α-(1→4) linked to d-GalpNAc. The roles of the Wzy polymerase and the four glycosyltransferases encoded by the KL89 gene cluster in the biosynthesis of the K89 CPS were assigned. Two glycosyltransferases, Gtr121 and Gtr122, link the d-Fucp3NAc to its neighboring sugars.

Capsule-Targeting Depolymerases Derived from Acinetobacter baumannii Prophage Regions.

In this study, several different depolymerases encoded in the prophage regions of Acinetobacter baumannii genomes have been bioinformatically predicted and recombinantly produced. The identified depolymerases possessed multi-domain structures and were identical or closely homologous to various proteins encoded in other A. baumannii genomes. This means that prophage-derived depolymerases are widespread, and different bacterial genomes can be the source of proteins with polysaccharide-degrading activities. For two depolymerases, the specificity to capsular polysaccharides (CPSs) of A. baumannii belonging to K1 and K92 capsular types (K types) was determined. The data obtained showed that the prophage-derived depolymerases were glycosidases that cleaved the A. baumannii CPSs by the hydrolytic mechanism to yield monomers and oligomers of the K units. The recombinant proteins with established enzymatic activity significantly reduced the mortality of Galleria mellonella larvae infected with A. baumannii of K1 and K92 capsular types. Therefore, these enzymes can be considered as suitable candidates for the development of new antibacterials against corresponding A. baumannii K types.