Function of bacteriophage G7C esterase tailspike in host cell adsorption.

Bacteriophages recognize and bind to their hosts with the help of receptor-binding proteins (RBPs) that emanate from the phage particle in the form of fibers or tailspikes. RBPs show a great variability in their shapes, sizes, and location on the particle. Some RBPs are known to depolymerize surface polysaccharides of the host while others show no enzymatic activity. Here we report that both RBPs of podovirus G7C - tailspikes gp63.1 and gp66 - are essential for infection of its natural host bacterium E. coli 4s that populates the equine intestinal tract. We characterize the structure and function of gp63.1 and show that unlike any previously described RPB, gp63.1 deacetylates surface polysaccharides of E. coli 4s leaving the backbone of the polysaccharide intact. We demonstrate that gp63.1 and gp66 form a stable complex, in which the N-terminal part of gp66 serves as an attachment site for gp63.1 and anchors the gp63.1-gp66 complex to the G7C tail. The esterase domain of gp63.1 as well as domains mediating the gp63.1-gp66 interaction is widespread among all three families of tailed bacteriophages.The work of the laboratory in the Winogradsky Institute was partially supported by Russian Science Foundation (RSF) grant #15–15‐0013

Protestant women in the late Soviet era: gender, authority, and dissent

At the peak of the anti-religious campaigns under Nikita Khrushchev, communist propaganda depicted women believers as either naïve dupes, tricked by the clergy, or as depraved fanatics; the Protestant “sektantka” (female sectarian) was a particularly prominent folk-devil. In fact, as this article shows, women’s position within Protestant communities was far more complex than either of these mythical figures would have one believe. The authors explore four important, but contested, female roles: women as leaders of worship, particularly in remote congregations where female believers vastly outnumbered their male counterparts; women as unofficial prophetesses, primarily within Pentecostal groups; women as mothers, replenishing congregations through high birth rates and commitment to their children’s religious upbringing; and women as political actors in the defence of religious rights. Using a wide range of sources, which include reports written by state officials, articles in the church journal, letters from church members to their ecclesiastical leaders in Moscow, samizdat texts, and oral history accounts, the authors probe women’s relationship with authority, in terms of both the authority of the (male) ministry within the church, and the authority of the Soviet state

Reprogramming bacteriophage host range: design principles and strategies for engineering receptor binding proteins

Bacteriophages (phages) use specialized tail machinery to deliver proteins and genetic material into a bacterial cell during infection. Attached at the distal ends of their tails are receptor binding proteins (RBPs) that recognize specific molecules exposed on host bacteria surfaces. Since the therapeutic capacity of naturally occurring phages is often limited by narrow host ranges, there is significant interest in expanding their host range via directed evolution or structure-guided engineering of their RBPs. Here, we describe the design principles of different RBP engineering platforms and draw attention to the mechanisms linking RBP binding and the correct spatial and temporal attachment of the phage to the bacterial surface. A deeper understanding of these mechanisms will directly benefit future engineering of more effective phage-based therapeutics. © 2021 ElsevierISSN:0958-1669ISSN:1879-042

Molecular Mechanisms of The Anomalous Thermal Aggregation of Green Fluorescent Protein

The peculiarities of thermal denaturation and interaction with water of the cycle-3 mutant of green fluorescent protein (GFP) were analyzed by NMR techniques and compared with those of bovine carbonic anhydrase II (BCA-II). Irreversible thermal denaturation was accompanied by massive GFP aggregation with no detectable accumulation of soluble denatured protein. Analysis of the spin diffusion data suggested that the internal part of the GFP β-can is involved in intensive interactions with water molecules. As a result, at high temperatures, the GFP structure does not unfold but rather breaks, consequently leading to enhanced protein aggregation. This is very different from typical BCA-II behavior

Genomic Sequencing and Biological Characteristics of a Novel Escherichia Coli Bacteriophage 9g, a Putative Representative of a New Siphoviridae Genus

Bacteriophage 9g was isolated from horse feces using Escherichia coli C600 as a host strain. Phage 9g has a slightly elongated capsid 62 × 76 nm in diameter and a non-contractile tail about 185 nm long. The complete genome sequence of this bacteriophage consists of 56,703 bp encoding 70 predicted open reading frames. The closest relative of phage 9g is phage PhiJL001 infecting marine alpha-proteobacterium associated with Ircinia strobilina sponge, sharing with phage 9g 51% of amino acid identity in the main capsid protein sequence. The DNA of 9g is resistant to most restriction endonucleases tested, indicating the presence of hypermodified bases. The gene cluster encoding a biosynthesis pathway similar to biosynthesis of the unusual nucleoside queuosine was detected in the phage 9g genome. The genomic map organization is somewhat similar to the typical temperate phage gene layout but no integrase gene was detected. Phage 9g efficiently forms stable associations with its host that continues to produce the phage over multiple passages, but the phage can be easily eliminated via viricide treatment indicating that no true lysogens are formed. Since the sequence, genomic organization and biological properties of bacteriophage 9g are clearly distinct from other known Enterobacteriaceae phages, we propose to consider it as the representative of a novel genus of the Siphoviridae family

Complete genome sequences of T5-related Escherichia coli bacteriophages DT57C and DT571/2 isolated from horse feces

We report the complete genome sequencing of two Escherichia coli T5-related bacteriophages, DT57C and DT571/2, isolated from the same specimen of horse feces. These two isolates share 96 % nucleotide sequence identity and can thus be considered representatives of the same novel species within the genus T5likevirus. The observed variation in the ltfA gene of these phages, resulting from a recent recombination event, may explain the observed host-range differences, suggesting that a modular mechanism makes a significant contribution to the short-term evolution (or adaptation) of T5-like phage genomes in the intestinal ecosystem. Comparison of our isolates to their closest relative, coliphage T5, revealed high overall synteny of the genomes and high conservation of the sequences of almost all structural proteins as well as of the other proteins with identified functions. At the same time, numerous alterations and non-orthologous replacements of non-structural protein genes (mostly of those with unknown functions) as well as substantial differences in tail fiber locus organization support the conclusion that DT57C and DT571/2 form a species-level group clearly distinct from bacteriophage T5

Branched Lateral Tail Fiber Organization in T5-Like Bacteriophages DT57C and DT571/2 is Revealed by Genetic and Functional Analysis

The T5-like siphoviruses DT57C and DT571/2, isolated from horse feces, are very closely related to each other, and most of their structural proteins are also nearly identical to T5 phage. Their LTFs (L-shaped tail fibers), however, are composed of two proteins, LtfA and LtfB, instead of the single Ltf of bacteriophage T5. In silico and mutant analysis suggests a possible branched structure of DT57C and DT571/2 LTFs, where the LtfB protein is connected to the phage tail via the LtfA protein and with both proteins carrying receptor recognition domains. Such adhesin arrangement has not been previously recognized in siphoviruses. The LtfA proteins of our phages are found to recognize different host O-antigen types: E. coli O22-like for DT57C phage and E. coli O87 for DT571/2. LtfB proteins are identical in both phages and recognize another host receptor, most probably lipopolysaccharide (LPS) of E. coli O81 type. In these two bacteriophages, LTF function is essential to penetrate the shield of the host's O-antigens. We also demonstrate that LTF-mediated adsorption becomes superfluous when the non-specific cell protection by O-antigen is missing, allowing the phages to bind directly to their common secondary receptor, the outer membrane protein BtuB. The LTF independent adsorption was also demonstrated on an O22-like host mutant missing O-antigen O-acetylation, thus showing the biological value of this O-antigen modification for cell protection against phages

Structure of the T4 baseplate and its function in triggering sheath contraction

Several systems, including contractile tail bacteriophages, the type VI secretion system and R-type pyocins, use a multiprotein tubular apparatus to attach to and penetrate host cell membranes. This macromolecular machine resembles a stretched, coiled spring (or sheath) wound around a rigid tube with a spike-shaped protein at its tip. A baseplate structure, which is arguably the most complex part of this assembly, relays the contraction signal to the sheath. Here we present the atomic structure of the approximately 6-megadalton bacteriophage T4 baseplate in its pre- and post-host attachment states and explain the events that lead to sheath contraction in atomic detail. We establish the identity and function of a minimal set of components that is conserved in all contractile injection systems and show that the triggering mechanism is universally conserved

Action of a minimal contractile bactericidal nanomachine.

R-type bacteriocins are minimal contractile nanomachines that hold promise as precision antibiotics1-4. Each bactericidal complex uses a collar to bridge a hollow tube with a contractile sheath loaded in a metastable state by a baseplate scaffold1,2. Fine-tuning of such nucleic acid-free protein machines for precision medicine calls for an atomic description of the entire complex and contraction mechanism, which is not available from baseplate structures of the (DNA-containing) T4 bacteriophage5. Here we report the atomic model of the complete R2 pyocin in its pre-contraction and post-contraction states, each containing 384 subunits of 11 unique atomic models of 10 gene products. Comparison of these structures suggests the following sequence of events during pyocin contraction: tail fibres trigger lateral dissociation of baseplate triplexes; the dissociation then initiates a cascade of events leading to sheath contraction; and this contraction converts chemical energy into mechanical force to drive the iron-tipped tube across the bacterial cell surface, killing the bacterium