Packing of Coat Protein Amphipathic and Transmembrane Helices in Filamentous Bacteriophage M13: Role of Small Residues in Protein Oligomeriza tion

Williams, Karen A.; Glibowicka, Mira; Li, Zuomei; Li, Hong; Khan, Amir R.; Chen, Yvonne M.Y.; Wang, Jing; Marvin, D.A.; Deber, Charles M.

doi:10.1006/jmbi.1995.0469

Cited by 74 publications

(55 citation statements)

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“…Y24 appears in various forms and is totally eliminated in X-2, a phage that shows a highly coiled character (43). Mutational studies and randomized mutagenesis (44,45) also point to the importance of the aromatic residues as well as several key hydrophobic residues. A simplified M13 coat protein obtained by alanine scanning mutagenesis (45) points to two key areas for phage assembly: N-terminal residues A7/9/10/18, F11, and L14, and the C-terminal residues I39, L41, F42, and F45.…”

Section: Figmentioning

confidence: 99%

The NMR–Rosetta capsid model of M13 bacteriophage reveals a quadrupled hydrophobic packing epitope

Morag

Sgourakis

Baker

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

115

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Filamentous phage are elongated semiflexible ssDNA viruses that infect bacteria. The M13 phage, belonging to the family inoviridae, has a length of ∼1 μm and a diameter of ∼7 nm. Here we present a structural model for the capsid of intact M13 bacteriophage using Rosetta model building guided by structure restraints obtained from magic-angle spinning solid-state NMR experimental data. The C5 subunit symmetry observed in fiber diffraction studies was enforced during model building. The structure consists of stacked pentamers with largely alpha helical subunits containing an N-terminal type II β-turn; there is a rise of 16.6-16.7 Å and a tilt of 36.1-36.6°between consecutive pentamers. The packing of the subunits is stabilized by a repeating hydrophobic stacking pocket; each subunit participates in four pockets by contributing different hydrophobic residues, which are spread along the subunit sequence. Our study provides, to our knowledge, the first magicangle spinning NMR structure of an intact filamentous virus capsid and further demonstrates the strength of this technique as a method of choice to study noncrystalline, high-molecular-weight molecular assemblies.solid-state NMR | magic-angle spinning | filamentous bacteriophage | structure determination | Rosetta modeling F ilamentous bacteriophage are long, thin, and semiflexible rod viruses that infect bacteria (1, 2). These large assemblies (∼15-35 MDa) contain a circular single-stranded (ss) DNA genome encapsulated in a protein shell. All filamentous phage have a similar life cycle and virion structure despite the relatively high number of strains, with DNA sequence homology varying from almost complete to very little. The unique phage properties make them ideal for a large range of applications such as phage display (3), DNA cloning and sequencing (4, 5), nanomaterial fabrication (6-8), and as drug-carrying nanomachines (9). In addition, filamentous viruses form a variety of liquid crystals driving the development of both theory and practice of softmatter physics (10, 11). Filamentous viruses are also associated with various diseases, e.g., CTXϕ phage in cholera toxin (12) and Pf4 phage in cystic fibrosis (13).Phage belonging to the Ff family (M13, fd, f1) are F-pilusspecific viruses that share almost identical genomes and very similar structures. M13 is a 16-MDa virus having a diameter of ∼7 nm and a length of ∼1 μm. The capsid is composed of several thousand identical copies of a major coat protein subunit arranged in a helical array surrounding a core of a circular ssDNA. The major coat proteins constitute ∼85% of the total virion mass, the ssDNA ∼12%, and all other minor proteins (gp3, gp6, gp7, gp9) that are specific for infection and assembly constitute about 3% of the total virion mass (1, 14).Previous structural models for a small number of phages have been obtained by means of X-ray fiber diffraction (15-19), static solid-state NMR (20, 21), and cryo-EM (22). Structural models for the Ff family have been proposed based on the three methods; however, ...

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Section: Figmentioning

confidence: 99%

The NMR–Rosetta capsid model of M13 bacteriophage reveals a quadrupled hydrophobic packing epitope

Morag

Sgourakis

Baker

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

115

View full text Add to dashboard Cite

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“…they cross. If the turn geometry is roughly the same for domain III of F-pilin then domains II and IV, as interacting ␣-helices, should each contain a patch of conserved amino acids that define interacting residues (see, for example, Williams et al, 1995). The mutagenesis experiments by which such residues can be identified have not been performed with F-pilin, but a comparison of related conjugative pilins serves the same purpose, at least to the extent of a preliminary test of the hypothesis.…”

Section: Structure Of Filament F-pilinmentioning

confidence: 99%

“…While the structural model elaborated here is consistent with many different experiments, it has yet to be explicitly tested. Saturation mutagenesis of F-pilin to identify residues important for filament assembly would provide one important test of the model (Williams et al, 1995), and other tests can readily be imagined. Finally, given the possiblity that the Ti plasmid virB 2 gene encodes a conjugative pilin related to F-pilin (Shirasu and Kado, 1993;Fullner et al, 1996), it is noteworthy that the VirB2 protein is identical to F-pilin at only 2 of the 15 positions in domains II, III, and IV at which F-pilin is identical to pEDP208 pilin (Shirasu and Kado, 1993) (Fig.…”

Section: Structure Of Filament F-pilinmentioning

confidence: 99%

Towards a structural biology of bacterial conjugation

Silverman

1997

Molecular Microbiology

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SummaryHorizontal DNA transfer among bacteria (conjugation) requires the formation of secure intercellular contacts before DNA transfer can occur. The formation of such contacts among Gram-negative bacteria is mediated by conjugative pili. Like other pili, conjugative pili are filaments extending from the surface of donor cells. They are composed, in so far as is known, entirely of conjugative pilin subunits. Here, we review the structure of F-pilin, the subunit of which F-pili are composed. We emphasize recent studies suggesting a specific domain organization for F-pilin and present a model of how these domains might be arranged in filament subunits.

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“…However, the mean (9, 2) values in model R Pf1 H over the region Ile22±Ala29 are (À63 AE 11, À40 AE 6 ), which are virtually identical to the mean over the whole -helix region (Table 2). It is doubtful that glycines are structurally signi®cant in ®lamentous bacteriophage, since glycines can be exchanged for other residues in the Ff structure (Williams et al, 1995). In particular, Gly24 of Pf1 corresponds in the aligned sequences to Gly23 of Ff (Nakashima et al, 1975;Marvin, 1990a,b) and Gly23 of Ff can be replaced by Arg, Asp, Ser or Ala while the phage remains viable (Williams et al, 1995).…”

mentioning

confidence: 99%

The molecular structure and structural transition of the α-helical capsid in filamentous bacteriophage Pf1

Welsh

Symmons

Marvin

2000

Acta Crystallogr D Biol Cryst

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The major coat protein in the capsid of Pf1 ®lamentous bacteriophage (Inovirus) forms a helical assembly of about 7000 identical protein subunits, each of which contains 46 amino-acid residues and can be closely approximated by a single gently curved -helix. Since the viral DNA occupies the core of the tubular capsid and appears to make no signi®cant speci®c interactions with the capsid proteins, the capsid is a simple model system for the study of the static and dynamic properties of -helix assembly. The capsid undergoes a reversible temperature-induced structural transition at about 283 K between two slightly different helix forms. The two forms can coexist without an intermediate state, consistent with a ®rst-order structural phase transition. The molecular model of the higher temperature form was re®ned using improved X-ray ®bre diffraction data and new re®nement and validation methods. The re®nement indicates that the two forms are related by a change in the orientation of the capsid subunits within the virion, without a signi®cant change in local conformation of the subunits. On the higher temperature diffraction pattern there is a region of observed intensity that is not consistent with a simple helix of identical subunits; it is proposed that the structure involves groups of three subunits which each have a slightly different orientation within the group. The grouping of subunits suggests that a change in subunit libration frequency could be the basis of the Pf1 structural transition; calculations from the model are used to explore this idea.

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Packing of Coat Protein Amphipathic and Transmembrane Helices in Filamentous Bacteriophage M13: Role of Small Residues in Protein Oligomeriza tion

Cited by 74 publications

References 0 publications

The NMR–Rosetta capsid model of M13 bacteriophage reveals a quadrupled hydrophobic packing epitope

The NMR–Rosetta capsid model of M13 bacteriophage reveals a quadrupled hydrophobic packing epitope

Towards a structural biology of bacterial conjugation

The molecular structure and structural transition of the α-helical capsid in filamentous bacteriophage Pf1

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