Random mutagenesis reveals a region important for gating of the yeast K+ channel Ykc1.
YKC1 (TOK1, DUK1, YORK) encodes the outwardly rectifying K+ channel of the yeast plasma membrane. Non-targeted mutations of YKC1 were isolated by their ability to completely block proliferation when expressed in yeast. All such mutations examined occurred near the cytoplasmic ends of the transmembrane segments following either of the duplicated P loops, which we termed the 'post-P loop' (PP) regions. These PP mutations specifically caused marked defects in the 'C1' states, a set of interrelated closed states that Ykc1 enters and exits at rates of tens to hundreds of milliseconds. These results indicate that the Ykc1 PP region plays a role in determining closed state conformations and that non-targeted mutagenesis and microbial selection can be a valuable tool for probing structure-function relationships of ion channels.
Loukin
S H
SH
Laboratory of Molecular Biology, University of Wisconsin, Madison 53706, USA.
Vaillant
B
B
Zhou
X L
XL
Spalding
E P
EP
Kung
C
C
Saimi
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015640:Ion Channel Gating; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009865:Oocytes; D018408:Patch-Clamp Techniques; D010957:Plasmids / Q000235:genetics; D015221:Potassium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013329:Structure-Activity Relationship; D014162:Transfection; D014981:Xenopus
D005656:Fungal Proteins; D015221:Potassium Channels; D029701:Saccharomyces cerevisiae Proteins; C095220:TOK1 protein, S cerevisiae