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Engineering of the Fluorescent‐Energy‐Conversion Arm of Phi29 DNA Packaging Motor for Single‐Molecule Studies
Author(s) -
Lee Tae Jin,
Zhang Hui,
Chang ChunLi,
Savran Cagri,
Guo Peixuan
Publication year - 2009
Publication title -
small
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.785
H-Index - 236
eISSN - 1613-6829
pISSN - 1613-6810
DOI - 10.1002/smll.200900467
Subject(s) - dna , green fluorescent protein , molecular motor , biophysics , atp hydrolysis , fluorescence , chemistry , biochemistry , nanotechnology , biology , materials science , physics , gene , enzyme , atpase , quantum mechanics
The bacteriophage phi29 DNA packaging motor contains a protein core with a central channel comprising twelve copies of re‐engineered gp10 protein geared by six copies of packaging RNA (pRNA) and a DNA packaging protein gp16 with unknown copies. Incorporation of this nanomotor into a nanodevice would be beneficial for many applications. To this end, extension and modification of the motor components are necessary for the linkage of this motor to other nanomachines. Here the re‐engineering of the motor DNA packaging protein gp16 by extending its length and doubling its size using a fusion protein technique is reported. The modified motor integrated with the eGFP‐gp16 maintains the ability to convert the chemical energy from adenosine triphosphate (ATP) hydrolysis to mechanical motion and package DNA. The resulting DNA‐filled capsid is subsequently converted into an infectious virion. The extended part of the gp16 arm is a fluorescent protein eGFP, which serves as a marker for tracking the motor in single‐molecule studies. The activity of the re‐engineered motor with eGFP‐gp16 is also observed directly with a bright‐field microscope via its ability to transport a 2‐µm‐sized cargo bound to the DNA.