The Journal of General Physiology, Vol 74, 457-478, Copyright © 1979 by The Rockefeller University Press
Modification of a voltage-gated K+ channel from sarcoplasmic reticulum by a pronase-derived specific endopeptidase
C Miller and RL Rosenberg
AK+ -selective membrane conductance channel from rabbit sarcoplasmic
reticulum (SR) is studied in an artificial planar phospholipid bilayer.
Membranes containing many such channels display voltage-dependent
conductance, which is well described by a two-state conformational
equilibrium with a free energy term linearly dependent on applied voltage.
Pronase-derived alkaline proteinase b (APb), when added to the side of the
membrane opposite to the SR vesicles (trans side), reduces the voltage
dependence of the K+ conductance. Single-channel fluctuation experiments
show that after APb treatment, the channel is still able to undergo
transitions between its open and closed states, but that the probability of
forming the open state is only slightly voltage-dependent. In terms of the
conformational model, the enzyme's primary effect is to reduce the
effective gating charge of the opening process by over 80%; a second effect
of APb is to reduce the internal free energy of opening from +1.2 to +0.4
kcal/mol. The kinetics of APb action are strongly voltage-dependent, so as
to indicate that the enzyme can react only with the channel's open state.
The results imply that the channel contains a highly charged polypeptide
region which moves in the direction perpendicular to the membrane plane
when transitions between the open and closed states occur. A lysine or
arginine residue in this region becomes exposed to the trans aqueous
solution when the channel is in its open conformation.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Facebook 
Reddit 
Technorati 
Twitter What's this?
