Functionalized Protein Nanotubes Based on the Bacteriophage vB_KleM-RaK2 Tail Sheath Protein.
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ABSTRACT:
SUBMITTER: Labutyte G
PROVIDER: S-EPMC8618960 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The contractile tail of bacteriophage T4 is a molecular machine that facilitates very high viral infection efficiency. Its major component is a tail sheath, which contracts during infection to less than half of its initial length. The sheath consists of 138 copies of the tail sheath protein, gene product (gp) 18, which surrounds the central non-contractile tail tube. The contraction of the sheath drives the tail tube through the outer membrane, creating a channel for the viral genome delivery. A crystal structure of about three quarters of gp18 has been determined and was fitted into cryo-electron microscopy reconstructions of the tail sheath before and after contraction. It was shown that during contraction, gp18 subunits slide over each other with no apparent change in their structure.
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Project description:Tail tubular protein A (TTPA) is a structural tail protein of Klebsiella pneumoniae bacteriophage KP32, and is responsible for adhering the bacteriophage to host cells. For the first time, we found that TTPA also exhibits lytic activity towards capsular exopolysaccharide (EPS) of the multiresistant clinical strain of Klebsiella pneumoniae, PCM2713, and thus should be regarded as a dual-function macromolecule that exhibits both structural and enzymatic actions. Here, we present our crystallographic and enzymatic studies of TTPA. TTPA was crystallized and X-ray diffraction data were collected to a resolution of 1.9 Å. In the crystal, TTPA molecules were found to adopt a tetrameric structure with α-helical domains on one side and β-strands and loops on the other. The novel crystal structure of TTPA resembles those of the bacteriophage T7 tail protein gp11 and gp4 of bacteriophage P22, but TTPA contains an additional antiparallel β-sheet carrying a lectin-like domain that could be responsible for EPS binding. The enzymatic activity of TTPA may reflect the presence of a peptidoglycan hydrolase domain in the α-helical region (amino acid residues 126 to 173). These novel results provide new insights into the enzymatic mechanism through which TTPA acts on polysaccharides.
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Project description:Bacteriophage phiKZ is a giant phage that infects Pseudomonas aeruginosa, a human pathogen. The phiKZ virion consists of a 1450 Å diameter icosahedral head and a 2000 Å-long contractile tail. The structure of the whole virus was previously reported, showing that its tail organization in the extended state is similar to the well-studied Myovirus bacteriophage T4 tail. The crystal structure of a tail sheath protein fragment of phiKZ was determined to 2.4 Å resolution. Furthermore, crystal structures of two prophage tail sheath proteins were determined to 1.9 and 3.3 Å resolution. Despite low sequence identity between these proteins, all of these structures have a similar fold. The crystal structure of the phiKZ tail sheath protein has been fitted into cryo-electron-microscopy reconstructions of the extended tail sheath and of a polysheath. The structural rearrangement of the phiKZ tail sheath contraction was found to be similar to that of phage T4.
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Project description:Achromobacter phage phiAxp-3, an N4-like bacteriophage, specifically recognize Achromobacter xylosoxidans lipopolysaccharide (LPS) as its receptor. PhiAxp-3 tail sheath protein (TSP, ORF69) shares 54% amino acid sequence identity with the TSP of phage N4 (gp65); the latter functions as a receptor binding protein and interacts with the outer membrane receptor NfrA of its host bacterium. Thus, we hypothesized that ORF69 is the receptor-binding protein of phiAxp-3. In the present study, a series of ORF69 truncation variants was constructed to identify the part(s) of this protein essential for binding to A. xylosoxidans LPS. Phage adsorption and enzyme-linked immunosorbent assay showed that amino acids 795-1195 of the TSP, i.e., ORF69(795-1195), are sufficient and essential for receptor and binding. The optimum temperature and pH for the functions of ORF69 and ORF69(795-1195) are 4/25°C and 7, respectively. In vitro cytotoxicity assays showed that ORF69 and ORF69(795-1195) were respectively toxic and non-toxic to a human immortalized normal hepatocyte cell line (LO2; doses: 0.375-12 ?g). The potential of this non-toxic truncated version of phiASP-3 TSP for clinical applications is discussed.
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Project description:Protein interactions in the assembly of the baseplate have been investigated. The baseplate of the phage T4 tail consists of a hub and six wedges which surround the former. Both reversible and irreversible interactions were found. Reversible association includes gp5 and gp27 (gp: gene product) which form a complex in a pH-dependent manner and gp18 polymerization, i.e. the tail sheath formation depends on the ionic strength. These reversible interactions were followed by irreversible or tight binding which pulls the whole association reaction to complete the assembly. The wedge assembly is strictly ordered which means that if one of the seven wedge proteins is missing, the assembly proceeds to that point and the remaining molecules stay non-associated. The strictly sequential assembly pathway is suggested to be materialized by successive conformational change upon binding, which can be shown by proteolytic probe.
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Project description:The majority of known bacteriophages have long tails that serve for bacterial target recognition and viral DNA delivery into the host. These structures form a tube from the viral capsid to the bacterial cell. The tube is formed primarily by a helical array of tail tube protein (TTP) subunits. In phages with a contractile tail, the TTP tube is surrounded by a sheath structure. Here, we report the first evidence that a phage TTP, gp17.1 of siphophage SPP1, self-assembles into long tubes in the absence of other viral proteins. gp17.1 does not exhibit a stable globular structure when monomeric in solution, even if it was confidently predicted to adopt the ?-sandwich fold of phage ? TTP. However, Fourier transform infrared and nuclear magnetic resonance spectroscopy analyses showed that its ?-sheet content increases significantly during tube assembly, suggesting that gp17.1 acquires a stable ?-sandwich fold only after self-assembly. EM analyses revealed that the tube is formed by hexameric rings stacked helicoidally with the same organization and helical parameters found for the tail of SPP1 virions. These parameters were used to build a pseudo-atomic model of the TTP tube. The large loop spanning residues 40-56 is located on the inner surface of the tube, at the interface between adjacent monomers and hexamers. In line with our structural predictions, deletion of this loop hinders gp17.1 tube assembly in vitro and interferes with SPP1 tail assembly during phage particle morphogenesis in bacteria.
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