A Single Atom Change Facilitates the Membrane Transport of Green Fluorescent Proteins in Mammalian Cells

Abstract

Direct delivery of proteins into mammalian cells is a challenging problem in biological and biomedical applications. The most common strategies for the delivery of proteins into the cells include the use of cell‐penetrating peptides or supercharged proteins. Herein, we show for the first time that a single atom change, hydrogen to halogen, at one of the tyrosine residues can increase the cellular entry of ∼28 kDa green fluorescent protein (GFP) in mammalian cells. The protein uptake is facilitated by a receptor‐mediated endocytosis and the cargo can be released effectively into cytosol by co‐treatment with the endosomolytic peptide ppTG21.

Single Small Molecule‐Assembled Mitochondria Targeting Nanofibers for Enhanced Photodynamic Cancer Therapy In Vivo

“…The results show the decrease in temperature inhibited cellular uptake of PQC NFs, which suggests the endocytosis PQC NFs is energy‐dependent (Figure 3c and Figure S12, Supporting information). [ 29 ] Four types of endocytosis inhibitors were then utilized to reveal the specific endocytic pathway of PQC NFs: sodium azide, chlorpromazine, genistein, and amiloride. Among these, both sodium azide and chlorpromazine decreased the cellular uptake of PQC NFs, which implicated that PQC NFs entered cells mainly through clathrin‐mediated endocytosis, rather than caveolae‐mediated endocytosis (genistein) or micropinocytosis (amiloride) (Figure 3d and Figure S12, Supporting information).…”

An Amphiphilic Sulfonatocalix[5]arene as an Activator for Membrane Transport of Lysine‐rich Peptides and Proteins

“…In the presence of sCx5‐6C, the cells showed distinct fluorescence, indicating uptake of FITC‐polyK (Figure 4 a). Notably, we still observed fluorescence when the cells were incubated at 4 °C (Figure 4 b), demonstrating that uptake can occur without energy [42] . A lower intensity was observed at 4 °C which can be explained by a slower diffusion of the sCx5‐6C/FITC‐polyK complex at this lower temperature.…”

“…Notably,w es till observed fluorescence when the cells were incubated at 4 8 8C ( Figure 4b), demonstrating that uptake can occur without energy. [42] A lower intensity was observed at 4 8 8Cw hich can be explained by aslower diffusion of the sCx5-6C/FITC-polyK complex at this lower temperature.H owever,w ec annot rule out that at 37 8 8Cthe energy dependent and independent processes occur simultaneously.T ransport, as signaled by intracellular fluorescence,w as not observed for the alternative activator candidates,n amely sCx4-5C and PyB (Figures 4c,d) and neither in the absence of any (Figure 4e). To further prove the entry of FITC-polyK activated by sCx5-6C,f low-cytometry assays were employed.…”

Recent Developments in Polymeric Assemblies and Functional Materials by Halogen Bonding

“…In most drug design, halogen atoms are incorporated to enhance the drug lipophilicity for improved penetration through lipid membranes. More studies on understanding their specific involvement in binding mechanism with artificial receptor models and membrane transport will lead to further progress in rational drug designing [128] . Needless to say that this applicability can pave the way for future design of self‐assembled biosensors for recognizing various biological processes and responding to different biological analytes [129] .…”