Alteration of channel activities and gating by mutations of slow ISK potassium channel.
ISK is a small membrane protein consisting of 129-130 amino acid residues with a single putative transmembrane domain and induces a very slow voltage-dependent K+ channel activity in the Xenopus oocyte expression system. We investigated the nature and structure-function relation of ISK by examining the effects of various mutations of ISK on the K+ channel activities measured in Xenopus oocytes. Deletion and truncation of the ISK protein indicated that the 63-amino acid sequence covering a transmembrane domain is sufficient for eliciting a K+ channel activity characteristic of ISK. Amino acid substitutions at a total of 31 positions within and surrounding the transmembrane domain caused different effects on the channel activity. A channel activity was enhanced by substitution of leucine with isoleucine at position 52 within the transmembrane domain, and the kinetic analysis of this mutation indicated that the enhancement of the channel activity is due to an alteration of a gating property of the ISK protein and thus supported the view that ISK forms an integral part of the K+ channel itself. The substitutions at many positions of the membrane-following region produced drastic reduction of the channel activity, and this is in marked contrast to the lack of effects of amino acid substitutions at the membrane-preceding region. Thus, the cytoplasmic portion immediately following the transmembrane domain plays a crucial role in inducing the channel activity of ISK.
Institute for Immunology, Kyoto University Faculty of Medicine, Japan.
T
T
Takumi
K
K
Moriyoshi
I
I
Aramori
T
T
Ishii
S
S
Oiki
Y
Y
Okada
H
H
Ohkubo
S
S
Nakanishi
D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000502:physiology; D002872:Chromosome Deletion; D006801:Humans; D015640:Ion Channel Gating; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology*; D010957:Plasmids; D011188:Potassium / Q000378:metabolism*; D015221:Potassium Channels / Q000235:genetics / Q000502:physiology*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014981:Xenopus
D015221:Potassium Channels; D012333:RNA, Messenger; D011188:Potassium