An engineered virus as a bright fluorescent tag and scaffold for cargo proteins--capture and transport by gliding microtubules.
We have demonstrated substantial capture and transport of fluorescently-labeled engineered cowpea mosaic virus (CPMV) using Drosophila kinesin-driven microtubules (MTs). The capture occurred through both NeutrAvidin (NA)-biotin and antibody (IgG)-antigen interactions. The MTs were derivatized with rabbit anti-chicken IgG or biotin, and the virus was conjugated with chicken IgG or NA. The CPMV conjugate was introduced into standard MT motility assays via convective flow at concentrations as high as 1.36 nM, and became bound to the MTs in densities as high as one virus per microm of MT length. When the CPMV conjugate was present at 17 pM, the average speed of the MTs bearing the NA-virus was 0.59 +/- 0.08 microm/sec, and that of those bearing IgG-virus was 0.52 +/- 0.15 microm/sec. These speeds are comparable to those of the unladen MTs (0.61 +/- 0.09 microm/sec), the presence of the virus on the MT causing only a small decrease in MT gliding speeds. The fluorescent CPMV appears to be superior to fluorescent polystyrene spheres of the same size, as both a reporter tag and a scaffold for MT-transported cargo proteins, because of its negligible non-specific adsorption and superior brightness. This work is important for the development of sensors based on nanolocomotion and biological recognition, or new strategies for the nanoassembly of biological structures.
Martin
Brett D
BD
Center for Bio/Molecular Science and Engineering, Code 6930, US Naval Research Laboratory, Washington, DC 20375, USA.
Soto
Carissa M
CM
Blum
Amy Szuchmacher
AS
Sapsford
K E
KE
Whitley
Jessica L
JL
Johnson
John E
JE
Chatterji
Anju
A
Ratna
Banahalli R
BR
D000327:Adsorption; D000818:Animals; D001360:Avidin / Q000737:chemistry*; D001709:Biotechnology / Q000379:methods; D001710:Biotin / Q000737:chemistry*; D002645:Chickens; D017798:Comovirus / Q000378:metabolism*; D004330:Drosophila / Q000378:metabolism*; D007074:Immunoglobulin G / Q000737:chemistry; D008301:Maleimides / Q000737:chemistry; D008870:Microtubules / Q000737:chemistry*; D053758:Nanoparticles / Q000737:chemistry; D036103:Nanotechnology / Q000379:methods*; D011817:Rabbits; D019809:Streptavidin / Q000737:chemistry
D007074:Immunoglobulin G; D008301:Maleimides; D001360:Avidin; C043592:maleimide; D001710:Biotin; D019809:Streptavidin