Abstract
Quantifying the passage of the large peptide protamine (Ptm) across CymA, a passive channel for cyclodextrin uptake,
is in the focus of this study. Using a reporter-pair based fluorescence membrane assay we detected the entry of Ptm into liposomes
containing CymA. The kinetics of the Ptm entry was independent of its concentration suggesting that the permeation across CymA
is the rate-limiting factor. Furthermore, we reconstituted single CymA channels into planar lipid bilayers and recorded the ion
current fluctuations in the presence of Ptm. To this end, we were able to resolve the voltage-dependent entry of single Ptm peptide
molecules into the channel. Extrapolation to zero voltage revealed about 1-2 events per second and long dwell times, in agreement
with the liposome study. Applied-field and steered molecular dynamics simulations provided an atomistic view on the permeation.
It can be concluded that a concentration gradient of 1 M Ptm leads to a translocation rate of about 1 molecule per second and
per channel.
Supplementary materials
Title
SI-25Dec2020
Description
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