Structure of the receptor-binding carboxy-terminal domain of bacteriophage T7 tail fibers

Abstract

The six bacteriophage T7 tail fibers, homo-trimers of gene product 17, are thought to be responsible for the first specific, albeit reversible, attachment to Escherichia coli lipopolysaccharide. The protein trimer forms kinked fibers comprised of an amino-terminal tail-attachment domain, a slender shaft, and a carboxyl-terminal domain composed of several nodules. Previously, we expressed, purified, and crystallized a carboxyl-terminal fragment comprising residues 371-553. Here, we report the structure of this protein trimer, solved using anomalous diffraction and refined at 2 Å resolution. Amino acids 371-447 form a tapered pyramid with a triangular cross-section composed of interlocked β-sheets from each of the three chains. The triangular pyramid domain has three α-helices at its narrow end, which are connected to a carboxyl-terminal three-blade β-propeller tip domain by flexible loops. The monomers of this tip domain each contain an eight-stranded β-sandwich. The exact topology of the β-sandwich fold is novel, but similar to that of knob domains of other viral fibers and the phage Sf6 needle. Several host-range change mutants have been mapped to loops located on the top of this tip domain, suggesting that this surface of the tip domain interacts with receptors on the cell surface.bacterial viruses | caudovirales | crystallography | infection | Podoviridae B acteriophages (bacterial viruses or phages) are important biological model systems and, because of the high specificity for their host bacteria, have found application in phage typing, food security, and phage therapy (1). Escherichia coli phage T7 is a member of the Podoviridae family of the Caudovirales (tailed phages) order (2). T7 is composed of an icosahedral capsid with a 20-nm short tail at one of the vertices (3, 4). The capsid is formed by the shell protein gene product (gp) 10 and encloses a DNA of 40 kb. A cylindrical structure composed of gp14, gp15, and gp16 is present inside the capsid (5), attached to the special vertex formed by the connector, a circular dodecamer of gp8 (6). Gp11 and gp12 form the tail; gp13, gp6.7, and gp7.3 have also been shown to be part of the virion and to be necessary for infection, although their location has not been established (7,8). Although extensive electron microscopy studies have been performed on phage T7 (3-6, 9), crystallographic studies have so far been limited to its nonstructural proteins.The main portion of the tail is composed of gp12, a large protein of which six copies are present (10); the small gp11 protein is also located in the tail (5). Attached to the tail are six fibers, each containing three copies of the gp17 protein. T7 tail fibers are elongated homo-trimers, which are responsible for initial, reversible, host cell recognition. A second, irreversible, decisionmaking interaction with the bacterial membrane is presumably mediated by one or more of the tail-tube proteins. DNA transfer into the host is then mediated by an extension formed by gp14-16 (7, 8, 11). Previously, we have reported t...

Outer Membrane Proteins Ail and OmpF of Yersinia pestis Are Involved in the Adsorption of T7-Related Bacteriophage Yep-phi

“…Considering that the initial binding of phage to bacteria is mediated by the interaction of its six gp17 tail fibers with bacterial surface receptors, gp17 protein of Yep-phi was expressed and purified as a His 6 fusion (11,34). Consistent with a previous report of T7 (34), we found that the purified recombinant gp17 protein of Yep-phi existed mainly as a multimer in solution. The multimer fraction was SDS resistant unless previously boiled (Fig.…”

“…2A). The alignment shows a highly conserved N-terminal moiety (amino acids 1 to 149), which belongs to domain family PHA00430 (34), indicating that they have the same mechanism of attaching the fiber to the phage as numerous T7-related phages. Aside from this region, the other sequence of the Yep-phi subgroup diversified from that of the other three (T7, T3, and PhiA1122).…”

Molecular architecture of tailed double-stranded DNA phages

“…For example, phage T7 has 6 tail fibers, 109,162,165 which interact with the 6-fold-symmetric and 12-fold symmetric tube proteins (Fig. 17B), whereas phage φ29 has 12 spindle-shaped tail spikes attached via the 12-foldsymmetric tube protein (Fig.…”

Conformational Changes Leading to T7 DNA Delivery upon Interaction with the Bacterial Receptor

“…The C-terminal portion of the fiber protein (gp17) is considered essential for virus-receptor interaction (12). To determine the structural component that interacts with rough LPS, we incubated rough LPS with recombinant fiber-less tail complexes and with fiber-containing tail complexes (13).…”

“…At least four proteins (gp8, gp11, gp12, and gp17) make up the T7 tail, which forms a central tubular structure with a closed, twisted nozzle at the bottom formed by a gp12 hexamer. This tubular assembly is surrounded by six protruding fibers specializing in receptor recognition, each of which is a gp17 trimer (12,13). Both proteins are docked to the connector (gp8) by a dodecameric ring termed adaptor or gatekeeper (gp11).…”