Supriyo Mukherjee, Kalyani Mahavidyalaya
The packaging of the viral genome is a fundamental process of the viral life cycle. Viruses are generally classified upon their genomic identity i.e., RNA or DNA as their genetic material. This affects the encapsidation strategy of the genetic material. In one encapsidation strategy, some viruses, especially most enveloped RNA viruses such as alphaviruses and flaviviruses, build a protein container called a capsid around the newly produced viral genome. In a second strategy, the most double-stranded DNA viruses, like poxvirus, create the capsid first and then fill the preformed capsid with the genetic material. We know that DNA is negatively charged and not so easy to pack together in a very small space due to repulsive force. The packaged viral DNA is usually in an extremely dense, nearly crystalline structure and about 60atm resultant pressure inside the viral capsid. Therefore, it requires a specialized DNA packaging motor for efficient packaging.
Powerful molecular motors like AAA+ motors, RecA- and FtsK-like ATPases, and ABC transporters are generally used by many viruses to encapsidate their viral DNA into procapsids during assembly, these motors are known as packaging ATPases, which converts chemical energy into mechanical energy using ATP hydrolysis.
In dsDNA viruses, the packaging machinery usually consists of a dodecameric portal protein at the 5-fold vertex and an ATP hydrolyzing pentameric motor protein which helps to package DNA. The dsDNA packaging motor is a transient component of the virus that does not remain with the virion. In a portal, the motor complex binds to the procapsids and dissociates on the completion of DNA packaging.
As per scientific research, viral DNA packaging motor has few types of mechanism. It is briefly discussed below-
Rotary motor mechanism – The dodecameric portal protein gp10 of bacteriophage phi29 is the first atomic structure of a packaging machine component. The portal has a wider end inside the capsid and a narrower end protruding from the capsid. The central channel is formed by alpha-helices and lined with negative charges, which gives a smooth passage of DNA. In this mechanism, the portal rotates by using the chemical energy from ATP hydrolysis which leads the DNA into the procapsids.
Linear motor mechanism (Based on electrostatic forces) – The viral dsDNA packaging motor protein generally has two different domains i.e., an N-terminal ATPase domain and a C-terminal domain that has nuclease activity. To retain packaging activity, these two domains need to be linked together physically. This linker consists of small amino acids. Recent studies on crystalline structures of the bacteriophage T4 packaging ATPase gp17 suggest that the N-terminal domain has a nucleotide-binding fold which is
structurally similar to monomeric helicases than hexameric ATPases. Conversely, the C-terminal domain belongs to the RnaseH/integrase superfamily. This mechanism was proposed for the bacteriophage T4 packaging motor, based on electrostatic interactions. A cis “arginine” finger in the N-terminal is positioned into the active center of ATPase that triggers ATP hydrolysis upon binding of dsDNA to the C-terminal domain of the gp17 subunit. The succeeding conformation change aligns the opposite charges in the N-and C-terminal domains, which lead to the pulling of the C-terminal domain towards the N-terminal domain, by the electrostatic force. This results in the packaging of two base pairs of dsDNA. The neighboring gp17 is now aligned with the dsDNA substrate for the forthcoming round of translocation because of the symmetry match between the five gp17s and the B-form DNA.
Non-integer step size motor mechanism – Although there is a plausible assumption that each motor subunit can package an integral number of base pairs for each hydrolyzed ATP molecule, scientific research reported a step size of 2.5 bps/ATP for the phi29 packaging motor. The phi29 procapsid and DNA are tethered to microbeads in two optical traps, by which the distance change between the two traps is measured to determine to package. There are two models, i.e., a piston-like model that assumes four of the subunits hydrolyze ATP while the fifth one retains it and it holds onto the DNA while reloading other subunits. This model requires that each motor subunit binds DNA in a non-specific manner. Conversely, The inchworm-like model proposes that only two subunits make contact with the DNA while conformational changes in all subunits create distortion which drives the packaging of 10bps.
Bacteriophage T4 genome packaging motor is the most powerful molecular motor known to date, which can generate a force of approx 60 piconewtons and package DNA at about 700bps.
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References:
- Joshua Pajak, Erik Dill, Emilio Reyes-Aldrete, Mark A White, Brian A Kelch, Paul J Jardine, Gaurav Arya, Marc C Morais, Atomistic basis of force generation, translocation, and coordination in a viral genome packaging motor, Nucleic Acids Research, 2021;, gkab372, https://doi.org/10.1093/nar/gkab372
- Simpson, A., Tao, Y., Leiman, P. et al. Structure of the bacteriophage φ29 DNA packaging motor. Nature 408, 745–750 (2000). https://doi.org/10.1038/3504712
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