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D A Marvin

Publications and source records attributed to D A Marvin.

At least 19 recordsLinked to original sources
PubMed1995

Packing of coat protein amphipathic and transmembrane helices in filamentous bacteriophage M13: role of small residues in protein oligomerization.

Filamentous bacteriophage M13, an important cloning and phage display vector, is encapsulated by ca 2700 copies of its 50-residue major coat protein (gene 8). This protein occurs as a membrane protein while stably inserted into its E. coli host inner membrane, and as a coat protein upon assembly and packing onto phage DNA in the lipid-free virion. To examine the specific protein-protein interactions underlying these processes, we used a combination of randomized and saturation mutagenesis of the entire gene 8 to assess the susceptibility of each position to mutation. In the resulting library of ca 100 viable M13 mutants, "small" residues (Ala,Gly,Ser), which constitute the non-polar face of the N-terminal amphipathic helical segment, and a face of the hydrophobic (effective transmembrane) helical segment, were found to be highly conserved. These results support a model in which coat protein packing is stabilized by the presence within each protein subunit of two "oligomerization segments", i.e. specific helical regions with faces rich in small residues which function to promote the close approach of alpha-helices.

Amino Acid Sequence↗
PubMed1995

Matching electrostatic charge between DNA and coat protein in filamentous bacteriophage. Fibre diffraction of charge-deletion mutants.

The virion of Ff (fd, f1, M13) filamentous bacteriophage consists of a long tube of coat protein subunits in a shingled, helical array, surrounding a genome of circular single-stranded DNA. Modified fd virions have been generated by a mutation (K48A) that removes one positive charge from each coat protein subunit in the C-terminal region of the polypeptide chain facing the DNA. The number of nucleotides in the mutant DNA is unchanged, but the K48A virions are 35% longer than wild-type. We have measured the X-ray diffraction attributable to single virions in hydrated gels of wild-type and K48A bacteriophages. Most of the diffraction pattern shows no significant difference between wild-type and K48A. Since the DNA is only about 12% by weight of the wild-type virion, the diffraction pattern is dominated by the protein contribution, and the absence of significant differences indicates that there are no significant changes in the symmetry or structure of the protein coat. But there is a change in the diffraction pattern in a region where the DNA and protein contributions are comparable. The diffraction pattern of the K48A mutant shows an increase in intensity of one of the weaker equatorial peaks, relative to wild-type, in a region where the protein contribution has negative sign but the DNA contribution has positive sign. This is consistent with a decrease in the ratio of DNA:protein per unit length of the K48A mutant. The results support the view that the protein forms a sheath lined with positive charges interacting electrostatically and non-specifically with a negatively charged DNA core of matching charge density. The lower positive charge density lining the capsid in the K48A mutant means that correspondingly fewer nucleotides can be packaged per coat protein subunit, which in turn requires an elongation of the DNA inside the virion. A longer virion is thus required to package the same amount of DNA. Within the error of measurement, the number of positive charges on the protein interacting with the DNA is the same in K48A as in the wild-type, despite the fact that the mutant is 35% longer than the wild-type.

Capsid↗
PubMed1990

Model-building studies of Inovirus: genetic variations on a geometric theme.

Inovirus (filamentous bacteriophage) is a simple system for studying the rules by which protein primary structure (amino acid sequence) controls secondary and higher order structure, and thereby function. The virus occurs naturally as a number of different strains with similar secondary and higher order structure, but the protein subunit that assembles to form the virion coat has quite different primary structures in different virus strains. Despite these differences in primary structure, the subunits of all strains have much the same size, about 50 residues, which are distributed by type in much the same way into three domains of primary structure: a collection of acidic residues in the N-terminal region, a hydrophobic domain of about 19 residues near the middle, and a collection of basic residues near the C-terminus. Each subunit can be closely approximated by an alpha-helix with its long axis roughly parallel to the fibre axis, sloping from large to small radius in the virion and interleaving between subunits in the next turn or level. The acidic residues near the N-terminus of the subunit face outwards on the virion surface, and explain the low isoelectric point of the virion; the basic residues near the C-terminus face inwards, where they neutralize the charge on the DNA at the core of the virion; and the hydrophobic central domain is involved in interactions which bind neighbouring subunits. Detailed X-ray fibre diffraction analysis of one strain gives the subunit structure. Comparative model-building studies of different strains illustrate the common structural principles.

Amino Acid Sequence↗
PubMed1989

Dynamics of telescoping Inovirus: a mechanism for assembly at membrane adhesions.

Telescoping of Inovirus (filamentous bacteriophage) into short hollow tubes by organic solvents suggests a molecular mechanism both for infection and for maturation of the virion at adhesions between the inner and outer bacterial membranes. The symmetry of alpha-helix subunit arrangement in the virion is related to the symmetry of leaf arrangement in plants (phyllotaxis) and is conserved in a molecular rearrangement model of the telescope.

Bacteriophages↗
PubMed1987

Pf1 Inovirus. Electron density distribution calculated by a maximum entropy algorithm from native fibre diffraction data to 3 A resolution and single isomorphous replacement data to 5 A resolution.

We have calculated the electron density distribution of the Pf1 strain of filamentous bacteriophage by a maximum entropy method. In the calculation we included native X-ray fibre diffraction data extending to 3 A resolution in the meridional direction on 60 layerlines that are resolved to 4 A in the equatorial direction, and lower resolution data from a single isomorphous derivative iodinated on the Tyr25 residue. The electron density map indicates that the 46-residue protein subunit is a single, gently curved stretch of alpha-helix with its axis at an angle of about 20 degrees to the axis of the virion. The alpha-helix subunit curves around the virion axis by about 1/6 turn, and decreases from about 27 A radius to about 13 A radius in the virion as the amino acid sequence of the subunit runs from the N terminus to the C terminus. Nearest-neighbour alpha-helical subunits are about 10 A apart along their length, and the axis of each subunit makes an unexpected negative angle with its nearest neighbours in the virion. To confirm the validity of the maximum entropy calculation, we have varied the constraints on the calculation. All variations result in either a map that is close to the original map or a map that cannot be interpreted in terms of secondary structure: we find only one map that makes structural sense.

Algorithms↗
PubMed1986

Structure of F-pili: reassessment of the symmetry.

Reassessment of the X-ray fibre diffraction patterns of F-pili using a more accurate subunit molecular weight suggests that subunits in F-pili are related by a fivefold rotation axis around the pilus axis. The identity of this fivefold symmetry with the fivefold rotation axis that relates the subunits in fd bacteriophage supports a simple model for tip-to-tip adsorption of bacteriophage to pili.

Bacterial Outer Membrane Proteins↗
PubMed1983

Pf1 bacteriophage replication--assembly complex. X-ray fibre diffraction of the high humidity form.

The helical intracellular nucleoprotein complex of Pf1 bacteriophage has been studied by X-ray fibre diffraction in various hydration states. The helix pitch changes from 44 A in dry fibres to 55 A in wet fibres, whereas the unit rise between subunits in the helix apparently does not change with humidity. This result indicates that the nucleoprotein assembly twists more readily than it stretches. This is consistent with its biological role of tightening the viral DNA into a more compact form for packaging in the virion.

Coliphages↗
PubMed1983

Proton currents and protein motion in membranes.

A mechanism is proposed whereby a proton gradient along a membrane-spanning alpha-helix is coupled to small changes in the torsional angles around the alpha-helix peptide bonds. Small concerted changes in the torsional angles are coupled to a change in the unit twist of the alpha-helix; a change in the unit twist is coupled to a change in the crossing angle between neighbouring alpha-helices; and a change in the crossing angle is coupled to a change in the size and shape of an assembly of alpha-helices. Following this logical linkage in one direction shows how a proton gradient could induce a pumping motion in an assembly of alpha-helices; following it in the other direction shows how motion in an assembly of alpha-helices could pump protons.

Biological Transport↗
PubMed1983

Maximum-entropy calculation of the electron density at 4 A resolution of Pf1 filamentous bacteriophage.

A 4 A electron-density map of Pf1 filamentous bacterial virus has been calculated from x-ray fiber diffraction data by using the maximum-entropy method. This method produces a map that is free of features due to noise in the data and enables incomplete isomorphous-derivative phase information to be supplemented by information about the nature of the solution. The map shows gently curved (banana-shaped) rods of density about 70 A long, oriented roughly parallel to the virion axis but slewing by about 1/6th turn while running from a radius of 28 A to one of 13 A. Within these rods, there is a helical periodicity with a pitch of 5 to 6 A. We interpret these rods to be the helical subunits of the virion. The position of strongly diffracted intensity on the x-ray fiber pattern shows that the basic helix of the virion is right handed and that neighboring nearly parallel protein helices cross one another in an unusual negative sense.

Bacteriophages↗
PubMed1981

Structure of the protein and DNA in fd filamentous bacterial virus.

The virion of filamentous bacterial viruses comprises a cylindrical protein shell of o.d. approximately 60 A and i.d. 20A, containing a single-stranded circular DNA molecule which has two oppositely directed but not base-paired strands extending the length of the virion. The assembly of the virion involves an intracellular prepackaging of the DNA with a viral DNA-binding protein which is then displaced by the coat protein as the growing virion crosses the bacterial membrane. Studies of the virion by X-ray fibre diffraction show that the protein coat consists largely of alpha-helices oriented roughly parallel to the axis of the virion. As the normal to a planar peptide tends to align normal to a magnetic field, it is possible to improve significantly the orientation of virions in fibres using a strong magnet. The success of this technique with the Pf1 strain of virus led us to apply it to the better-known fd (f1, M13) strain. We report here new information about the arrangement of protein and DNA in the fd virion obtained from the improved diffraction pattern (Fig. 1).

Bacteriophages↗