Compared to the conventional approach to protein delivery (top), the Gao team’s simple new tagging system (bottom) bypasses endocytosis, enabling more proteins to reach the inner cell, which allows real-time imaging and makes treatments more effective. Credit: Gao Lab
Calling it a “plug-and-play platform,” the authors note the system is especially suited to small proteins, such as antibody fragments, nanobodies, engineered antibodies called single-chain variable fragments (scFvs), peptides, and some enzymes.
To make the protein delivery tag, Dr. Gao and his co-authors, Wanyi Tai and Pengfei Zhao, research scientists in Dr. Gao’s lab, link two copies of cholesterol, a natural component of cell membranes that moves easily through cell walls, to a common biocompatible blue dye.
When they mix the tag with proteins, the dye end of the tag latches on to the protein molecules, and the exposed cholesterol makes the cargo protein compatible with the cell membrane lipid bilayers, the authors suggest.
The authors hypothesize that because the protein tag is formed with a slightly weaker, noncovalent bond, the protein molecule is able to separate in the membrane bilayer, where its incompatibility forces it either into the cell’s center or back out.
The researchers tested their technology in 10 types of human cells, using a range of protein sizes. They found that direct permeation into the cells was most effective with smaller proteins, likely due to the larger molecules causing a bulge in the cell membrane.
“This simple yet powerful technology not only enables immuno-labeling of intracellular targets in live cells for dynamic imaging, but also opens exciting opportunities for the development of protein therapeutics and cell engineering,” the authors write.
Cytosolic delivery of proteins by cholesterol tagging, W. Tai et al., Science Advances 2020