A thermophilic phage uses a small terminase protein with a fixed helix-turn-helix geometry

SUMMARYTailed bacteriophage use a DNA packaging motor to encapsulate their genome during viral particle assembly. The small terminase (TerS) component acts as a molecular matchmaker by recognizing the viral genome as well as the main motor component, the large terminase (TerL). How TerS binds DNA and the TerL protein remains unclear. Here, we identify the TerS protein of the thermophilic bacteriophage P74-26. TerSP76-26 oligomerizes into a nonamer that binds DNA, stimulates TerL ATPase activity, and inhibits TerL nuclease activity. Our cryo-EM structure shows that TerSP76-26 forms a ring with a wide central pore and radially arrayed helix-turn-helix (HTH) domains. These HTH domains, which are thought to bind DNA by wrapping the helix around the ring, are rigidly held in an orientation distinct from that seen in other TerS proteins. This rigid arrangement of the putative DNA binding domain imposes strong constraints on how TerSP76-26 can bind DNA. Finally, the TerSP76-26 structure lacks the conserved C-terminal β-barrel domain used by other TerS proteins for binding TerL, suggesting that a well-ordered C-terminal β-barrel domain is not necessary for TerS to carry out its function as a matchmaker.

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A thermophilic phage uses a small terminase protein with a fixed helix–turn–helix geometry

Journal of Biological Chemistry ◽

10.1074/jbc.ra119.012224 ◽

2020 ◽

Vol 295 (12) ◽

pp. 3783-3793 ◽

Cited By ~ 1

Author(s):

Janelle A. Hayes ◽

Brendan J. Hilbert ◽

Christl Gaubitz ◽

Nicholas P. Stone ◽

Brian A. Kelch

Keyword(s):

Double Helix ◽

Nuclease Activity ◽

Dna Packaging ◽

Particle Assembly ◽

Dna Packaging Motor ◽

Motor Component ◽

Thermophilic Bacteriophage ◽

Viral Maturation ◽

Viral Particle Assembly

Tailed bacteriophages use a DNA-packaging motor to encapsulate their genome during viral particle assembly. The small terminase (TerS) component of this DNA-packaging machinery acts as a molecular matchmaker that recognizes both the viral genome and the main motor component, the large terminase (TerL). However, how TerS binds DNA and the TerL protein remains unclear. Here we identified gp83 of the thermophilic bacteriophage P74-26 as the TerS protein. We found that TerSP76-26 oligomerizes into a nonamer that binds DNA, stimulates TerL ATPase activity, and inhibits TerL nuclease activity. A cryo-EM structure of TerSP76-26 revealed that it forms a ring with a wide central pore and radially arrayed helix–turn–helix domains. The structure further showed that these helix–turn–helix domains, which are thought to bind DNA by wrapping the double helix around the ring, are rigidly held in an orientation distinct from that seen in other TerS proteins. This rigid arrangement of the putative DNA-binding domain imposed strong constraints on how TerSP76-26 can bind DNA. Finally, the TerSP76-26 structure lacked the conserved C-terminal β-barrel domain used by other TerS proteins for binding TerL. This suggests that a well-ordered C-terminal β-barrel domain is not required for TerSP76-26 to carry out its matchmaking function. Our work highlights a thermophilic system for studying the role of small terminase proteins in viral maturation and presents the structure of TerSP76-26, revealing key differences between this thermophilic phage and its mesophilic counterparts.

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Are phytoplankton population density maxima predictable through analysis of host and viral genomic DNA content?

Journal of the Marine Biological Association of the United Kingdom ◽

10.1017/s0025315406013397 ◽

2006 ◽

Vol 86 (3) ◽

pp. 491-498 ◽

Cited By ~ 27

Author(s):

Chris M. Brown ◽

Janice E. Lawrence ◽

Douglas A. Campbell

Keyword(s):

Population Density ◽

Dna Content ◽

Viral Genome ◽

Particle Assembly ◽

Phytoplankton Population ◽

Strong Linear Correlation ◽

Viral Progeny ◽

A Cell ◽

Viral Proliferation ◽

Viral Particle Assembly

Phytoplankton:virus interactions are important factors in aquatic nutrient cycling and community succession. The number of viral progeny resulting from an infection of a cell critically influences the propagation of infection and concomitantly the dynamics of phytoplankton populations. Host nucleotide content may be the resource limiting viral particle assembly. We present evidence for a strong linear correlation between measured viral burst sizes and viral burst sizes predicted from the host DNA content divided by the viral genome size, across a diversity of phytoplankton:viral pairs. An analysis of genome sizes therefore supports predictions of taxon-specific phytoplankton population density thresholds beyond which viral proliferation can trim populations or terminate phytoplankton blooms. We present corollaries showing that host:virus interactions may place evolutionary pressure towards genome reduction of both phytoplankton hosts and their viruses.

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The large terminase DNA packaging motor grips DNA with its ATPase domain for cleavage by the flexible nuclease domain

Nucleic Acids Research ◽

10.1093/nar/gkw1356 ◽

2017 ◽

pp. gkw1356 ◽

Cited By ~ 5

Author(s):

Brendan J. Hilbert ◽

Janelle A. Hayes ◽

Nicholas P. Stone ◽

Rui-Gang Xu ◽

Brian A. Kelch

Keyword(s):

Dna Packaging ◽

Atpase Domain ◽

Nuclease Domain ◽

Dna Packaging Motor ◽

Large Terminase

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A viral genome packaging ring-ATPase is a flexibly coordinated pentamer

10.1101/2021.03.08.434477 ◽

2021 ◽

Author(s):

Li Dai ◽

Digvijay Singh ◽

Suoang Lu ◽

Vishal Kottadiel ◽

Reza Vafabakhsh ◽

...

Keyword(s):

Single Molecule ◽

Viral Genome ◽

Motor Coordination ◽

Bacteriophage T4 ◽

Dna Packaging ◽

Biological Functions ◽

Dna Translocation ◽

Genome Packaging ◽

Dna Packaging Motor ◽

Direct Counting

Multi-subunit ring-ATPases carry out a myriad of biological functions, including genome packaging in viruses. Though the basic structures and functions of these motors have been well-established, the mechanisms of ATPase firing and motor coordination are poorly understood. Here, by direct counting using single-molecule fluorescence, we have determined that the active bacteriophage T4 DNA packaging motor consists of five subunits of gp17. By systematically doping motors with an ATPase-defective subunit and selecting single motors containing a precise count of active/inactive subunit(s), we found, unexpectedly, that the packaging motor can tolerate an inactive sub-unit. However, motors containing an inactive subunit(s) exhibit fewer DNA engagements, a higher failure rate in encapsidation, reduced packaging velocity, and increased pausing. These findings suggest a new packaging model in which the motor, by re-adjusting its grip on DNA, can skip an inactive subunit and resume DNA translocation, contrary to the prevailing notion of strict coordination amongst motor subunits of other packaging motors.

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A viral genome packaging ring-ATPase is a flexibly coordinated pentamer

Nature Communications ◽

10.1038/s41467-021-26800-z ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Li Dai ◽

Digvijay Singh ◽

Suoang Lu ◽

Vishal I. Kottadiel ◽

Reza Vafabakhsh ◽

...

Keyword(s):

Single Molecule ◽

Viral Genome ◽

Motor Coordination ◽

Bacteriophage T4 ◽

Dna Packaging ◽

Biological Functions ◽

Dna Translocation ◽

Single Molecule Fluorescence ◽

Genome Packaging ◽

Dna Packaging Motor

AbstractMulti-subunit ring-ATPases carry out a myriad of biological functions, including genome packaging in viruses. Though the basic structures and functions of these motors have been well-established, the mechanisms of ATPase firing and motor coordination are poorly understood. Here, using single-molecule fluorescence, we determine that the active bacteriophage T4 DNA packaging motor consists of five subunits of gp17. By systematically doping motors with an ATPase-defective subunit and selecting single motors containing a precise number of active or inactive subunits, we find that the packaging motor can tolerate an inactive subunit. However, motors containing one or more inactive subunits exhibit fewer DNA engagements, a higher failure rate in encapsidation, reduced packaging velocity, and increased pausing. These findings suggest a DNA packaging model in which the motor, by re-adjusting its grip on DNA, can skip an inactive subunit and resume DNA translocation, suggesting that strict coordination amongst motor subunits of packaging motors is not crucial for function.

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