Effects of Portal Protein Primary Structure Mutations on Viral Genomic Packaging Capabilities

The process by which bacteriophages package and maintain double-stranded DNA within the protein capsid has been subject to a great deal of research in recent years. Given the increased interest in particular fields of medicine, such as gene therapy, the desire to understand how the DNA packaging motor in the bacteriophages operates has opened many doors for revolutionary scientific research in the fields of biology, chemistry, biophysics and nanotechnology. Due to the universality of the structure of bacteriophages, the study of one particular bacteriophage could provide insight into other types of bacteriophages. As such, the bacteriophage Phi29 was utilized, and its DNA packaging motor is composed of a portal protein, packaging RNA and a viral ATPase. The focus of the research is on the portal protein that is a channel protein in the capsid of the bacteriophage through which the genome DNA can translocate during the DNA packaging. Many previous studies has indicated that the portal protein may not only serve as a passive conduit for the genome DNA but also play other critical roles during the packaging process. However, they are still intensively debated and discussed today. A recent study in our lab has found the portal protein may serve a Flashing Brownian ratchet in the packaging process, which can assist the packaging of the genome DNA against its concentration gradient across the portal protein at the late stage of DNA packaging. In order to study the relationship between the structure and the function of the portal proteins, we performed planar bilayer membrane measurements for mutant portal proteins. Within this experiment, we were able to establish a potential gradient across the membrane and use this potential to establish an ion selectivity for the membrane. From this point, the abilities of the portal protein to package an anionic genome were inferred. Our results have shown that the charges of the inner-ring amino acids alter the ion-selectivity of the channel which is pivotal to understanding the impacts of charges of amino acid residues on the genome packaging efficiency.http://opus.ipfw.edu/stu_symp2017/1004/thumbnail.jp