Function of a viral genome packaging motor from bacteriophage T4 is insensitive to DNA sequence

2021 ◽  
Author(s):  
Douglas E. Smith ◽  
Youbin E. Mo ◽  
Nick Keller ◽  
Damian delToro ◽  
Neeti Ananthaswamy ◽  
...  
Keyword(s):  
Dna Sequence ◽  
Viral Genome ◽  
Bacteriophage T4 ◽  
Genome Packaging

scholarly journals Function of a viral genome packaging motor from bacteriophage T4 is insensitive to DNA sequence

2020 ◽  
Vol 48 (20) ◽  
pp. 11602-11614
Author(s):  
Youbin Mo ◽  
Nicholas Keller ◽  
Damian delToro ◽  
Neeti Ananthaswamy ◽  
Stephen C Harvey ◽  
...  
Keyword(s):  
Dna Sequence ◽  
Motor Function ◽  
Optical Tweezers ◽  
Single Molecule ◽  
Dna Sequences ◽  
Viral Genome ◽  
Bacteriophage T4 ◽  
Repeated Measurements ◽  
Genome Packaging ◽  
Sequence Dependence

Abstract Many viruses employ ATP-powered motors during assembly to translocate DNA into procapsid shells. Previous reports raise the question if motor function is modulated by substrate DNA sequence: (i) the phage T4 motor exhibits large translocation rate fluctuations and pauses and slips; (ii) evidence suggests that the phage phi29 motor contacts DNA bases during translocation; and (iii) one theoretical model, the ‘B-A scrunchworm’, predicts that ‘A-philic’ sequences that transition more easily to A-form would alter motor function. Here, we use single-molecule optical tweezers measurements to compare translocation of phage, plasmid, and synthetic A-philic, GC rich sequences by the T4 motor. We observed no significant differences in motor velocities, even with A-philic sequences predicted to show higher translocation rate at high applied force. We also observed no significant changes in motor pausing and only modest changes in slipping. To more generally test for sequence dependence, we conducted correlation analyses across pairs of packaging events. No significant correlations in packaging rate, pausing or slipping versus sequence position were detected across repeated measurements with several different DNA sequences. These studies suggest that viral genome packaging is insensitive to DNA sequence and fluctuations in packaging motor velocity, pausing and slipping are primarily stochastic temporal events.

scholarly journals Function of a Viral Genome Packaging Motor from Bacteriophage T4 is Insensitive to DNA Sequence

2021 ◽  
Vol 120 (3) ◽  
pp. 36a
Author(s):  
Douglas E. Smith ◽  
Youbin Mo ◽  
Nick Keller ◽  
Damian delToro ◽  
Neeti Ananthaswamy ◽  
...  
Keyword(s):  
Dna Sequence ◽  
Viral Genome ◽  
Bacteriophage T4 ◽  
Genome Packaging

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

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.

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

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.

scholarly journals Viral genome packaging terminase cleaves DNA using the canonical RuvC-like two-metal catalysis mechanism

2017 ◽  
pp. gkw1354 ◽  
Author(s):  
Rui-Gang Xu ◽  
Huw T. Jenkins ◽  
Maria Chechik ◽  
Elena V. Blagova ◽  
Anna Lopatina ◽  
...  
Keyword(s):  
Viral Genome ◽  
Genome Packaging ◽  
Metal Catalysis ◽  
Catalysis Mechanism

scholarly journals Insights into Specific DNA Recognition during the Assembly of a Viral Genome Packaging Machine

2002 ◽  
Vol 9 (5) ◽  
pp. 981-991 ◽  
Author(s):  
Tonny de Beer ◽  
Jenny Fang ◽  
Marcos Ortega ◽  
Qin Yang ◽  
Levi Maes ◽  
...  
Keyword(s):  
Viral Genome ◽  
Dna Recognition ◽  
Genome Packaging ◽  
Packaging Machine

scholarly journals Energy-Independent Helicase Activity of a Viral Genome Packaging Motor

Biochemistry ◽  
2011 ◽  
Vol 51 (1) ◽  
pp. 391-400 ◽  
Author(s):  
Jenny R. Chang ◽  
Benjamin T. Andrews ◽  
Carlos E. Catalano
Keyword(s):  
Viral Genome ◽  
Helicase Activity ◽  
Genome Packaging
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