The effect of N- or C-terminal alterations of the connector of bacteriophage phi29 DNA packaging motor on procapsid assembly, pRNA binding, and DNA packaging.
Double-stranded DNA viruses package their genomes into procapsids via an ATP-driven nanomotor. This ingenious motor configuration has inspired the development of biomimetics in nanotechnology. Bacteriophage varphi29 DNA-packaging motor has been a popular tool in nanomedicine. To provide information for further motor modification, conjugation, labeling, and manufacturing, the connector protein gp10 of the varphi29 DNA packaging motor was truncated, mutated, and extended. A 25-residue deletion or a 14-residue extension at the C terminus of gp10 did not affect procapsid assembly. A 42-amino acid extension at the N terminus did not interfere with the procapsid assembly but significantly decreased the DNA-packaging efficiency. DNA-packaging activity was restored upon protease cleavage of the extended region. Replacing the N-terminal peptide containing arginine and lysine with a histidine-rich peptide did not affect procapsid assembly but completely inhibited the packaging RNA (pRNA) binding to the connector and hindered subsequent DNA packaging. These results indicate that (1) the N-terminal arginine-lysine residues play a critical role in pRNA binding but are not essential for procapsid assembly; (2) the connector core, but not the flexible N- or C-terminal domains, is responsible for signaling the procapsid assembly; (3) pRNA binds to the connector as a result of electrostatic interactions between the polyanionic nature of nucleic acids and the cationic side groups of the amino acids, similar to RNA binding to Tat or polyArg.
Cai
Ying
Y
Department of Biomedical Engineering, The Vontz Center for Molecular Studies, College of Engineering and College of Medicine, University of Cincinnati, Cincinnati, Ohio 45267, USA.
Xiao
Feng
F
Guo
Peixuan
P
D017103:Bacillus Phages / Q000235:genetics*; D002213:Capsid; D042003:DNA Packaging; D004279:DNA, Viral / Q000235:genetics*; D046529:Microscopy, Electron, Transmission; D010957:Plasmids; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism*
D004279:DNA, Viral; D012367:RNA, Viral