scholarly journals Designing a nine cysteine-less DNA packaging motor from bacteriophage T4 reveals new insights into ATPase structure and function

Virology ◽  
2014 ◽  
Vol 468-470 ◽  
pp. 660-668 ◽  
Author(s):  
Kiran Kondabagil ◽  
Li Dai ◽  
Reza Vafabakhsh ◽  
Taekjip Ha ◽  
Bonnie Draper ◽  
...  
Keyword(s):  
Bacteriophage T4 ◽  
Dna Packaging ◽  
Structure And Function ◽  
Dna Packaging Motor ◽  
And Function

VERSATILE DNA-PACKAGING NANOMOTOR OF BACTERIOPHAGE phi29 WITH APPLICATIONS IN NANOBIOTECHNOLOGY

Nano LIFE ◽  
2010 ◽  
Vol 01 (01n02) ◽  
pp. 45-62 ◽  
Author(s):  
JIA GENG ◽  
ANNE P. VONDERHEIDE
Keyword(s):  
Viral Assembly ◽  
Dna Packaging ◽  
Structure And Function ◽  
Bacteriophage Phi29 ◽  
Nanosized Materials ◽  
Dna Packaging Motor ◽  
Electrophysiological Measurement ◽  
And Function

Nanobiotechnology entails the use of nanosized materials to build structures that can be applied in both biotechnology and medicine. In one vein of this field of study, scientists seek to mimic the wide variety of nanomachines and macromolecular structures that exist in nature and to replicate them in both structure and function. As a most intriguing example, the bacterial virus phi29 uses a self-contained nanomotor to package its DNA after replication. The 30-nm nanomotor contains 12 copies of a protein (gp10) which together form a 3.6-nm central channel through which the genomic DNA passes into the procapsid during viral assembly and exits during infection. This connector has been recently reengineered and embedded into a lipid bilayer, creating a system with tremendous application for DNA detection and characterization through electrophysiological measurement. A second component of the phi29 bacteriophage is an ATP-binding pRNA that forms a hexameric ring to gear the motor. The pRNA has been utilized to construct nanoparticles of dimers, trimers, hexamers and patterned superstructures via the interaction of two interlocking loops. Such structures constructed via bottom-up assembly have been used in the delivery of drugs, siRNA, ribozymes, and genes to specific cells, both in vitro and in vivo. This review summarizes current studies on the structure, function, and mechanism of the phi29 DNA packaging motor, as well as addresses the applications of these motor components in the field of nanobiotechnology.

scholarly journals Structure and function of the small terminase component of the DNA packaging machine in T4-like bacteriophages

2011 ◽  
Vol 109 (3) ◽  
pp. 817-822 ◽  
Author(s):  
S. Sun ◽  
S. Gao ◽  
K. Kondabagil ◽  
Y. Xiang ◽  
M. G. Rossmann ◽  
...  
Keyword(s):  
Dna Packaging ◽  
Structure And Function ◽  
Packaging Machine ◽  
And Function

scholarly journals Mechanism of Coordination of the Bacteriophage T4 DNA Packaging Motor Analyzed by Real-Time Single Molecule Fluorescence Assay

2016 ◽  
Vol 110 (3) ◽  
pp. 46a ◽  
Author(s):  
Li Dai ◽  
Digvijay Singh ◽  
Reza Vafabakhsh ◽  
Marthandan Mahalingam ◽  
Vishal Kottadiel ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Bacteriophage T4 ◽  
Dna Packaging ◽  
Fluorescence Assay ◽  
Single Molecule Fluorescence ◽  
Dna Packaging Motor

scholarly journals Altering the speed of a DNA packaging motor from bacteriophage T4

2017 ◽  
Vol 45 (19) ◽  
pp. 11437-11448 ◽  
Author(s):  
Siying Lin ◽  
Tanfis I. Alam ◽  
Vishal I. Kottadiel ◽  
Carl J. VanGessel ◽  
Wei-Chun Tang ◽  
...  
Keyword(s):  
Bacteriophage T4 ◽  
Dna Packaging ◽  
Dna Packaging Motor

scholarly journals Structure and Function of the T4 Spackle Protein Gp61.3

Viruses ◽  
2020 ◽  
Vol 12 (10) ◽  
pp. 1070
Author(s):  
Shuji Kanamaru ◽  
Kazuya Uchida ◽  
Mai Nemoto ◽  
Alec Fraser ◽  
Fumio Arisaka ◽  
...  
Keyword(s):  
Phage Particle ◽  
Bacteriophage T4 ◽  
Structure And Function ◽  
Lysozyme Activity ◽  
T4 Lysozyme ◽  
Infection Cycle ◽  
Active Site Cleft ◽  
Targeting Peptide ◽  
And Function ◽  
Peptidoglycan Layer

The bacteriophage T4 genome contains two genes that code for proteins with lysozyme activity—e and 5. Gene e encodes the well-known T4 lysozyme (commonly called T4L) that functions to break the peptidoglycan layer late in the infection cycle, which is required for liberating newly assembled phage progeny. Gene product 5 (gp5) is the tail-associated lysozyme, a component of the phage particle. It forms a spike at the tip of the tail tube and functions to pierce the outer membrane of the Escherichia coli host cell after the phage has attached to the cell surface. Gp5 contains a T4L-like lysozyme domain that locally digests the peptidoglycan layer upon infection. The T4 Spackle protein (encoded by gene 61.3) has been thought to play a role in the inhibition of gp5 lysozyme activity and, as a consequence, in making cells infected by bacteriophage T4 resistant to later infection by T4 and closely related phages. Here we show that (1) gp61.3 is secreted into the periplasm where its N-terminal periplasm-targeting peptide is cleaved off; (2) gp61.3 forms a 1:1 complex with the lysozyme domain of gp5 (gp5Lys); (3) gp61.3 selectively inhibits the activity of gp5, but not that of T4L; (4) overexpression of gp5 causes cell lysis. We also report a crystal structure of the gp61.3-gp5Lys complex that demonstrates that unlike other known lysozyme inhibitors, gp61.3 does not interact with the active site cleft. Instead, it forms a “wall” that blocks access of an extended polysaccharide substrate to the cleft and, possibly, locks the enzyme in an “open-jaw”-like conformation making catalysis impossible.

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.

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