Characterization of Gating and Peptide Block of mSlo, a Cloned Calcium-
Dependent Potassium Channel.
Deirdre A. Sullivan, Mats Holmqvist and Irwin B. Levitan.
Department of Biochemistry and Volen Center for Complex Systems, Brandeis
University, Waltham, MA 02254.
APStracts 4:182N, 1997.
ABSTRACT
The twenty amino acid Shaker inactivation peptide blocks mSlo, a cloned
calcium-dependent potassium channel. Changing the charge and degree of
hydrophobicity of the peptide alters its blocking kinetics. A "triple mutant"
mSlo channel was constructed in which three amino acids (T256, S259, and
L262)_, equivalent to those identified as part of the peptide's receptor site
in the S4-S5 cytoplasmic loop region_ of the Shaker channel, were mutated
_simultaneously to alanines. These mutations produce only limited changes in
the channel's susceptibility to block by a series of peptides of varying
charge and hydrophobicity, but do alter channel gating. The "triple mutant"
channel shows a significant shift in its calcium-activation curve, as compared
to the wild type channel._ Analysis of the corresponding single amino acid
mutations shows that mutation at position L262 causes the most dramatic change
in mSlo gating. These results suggest that the three amino acids mutated in
the mSlo S4-S5 loop may contribute to, but are not essential for, peptide
binding. On the other hand they do play a critical_ role in the channel's
calcium-sensing mechanism._
Received 23 May 1997; accepted in final form 1 August 1997.
APS Manuscript Number J433-7.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1997 The American Physiological Society.
Published in APStracts on 28 August 1997