On the mechanism of a mutated and abnormally functioning gamma-aminobutyric acid (A) receptor linked to epilepsy.
A recent report indicates that a lysine-to-methionine mutation (K289M) in the gamma2 subunit of a human gamma-aminobutyric acid neurotransmitter receptor, the GABA(A) receptor, is linked to generalized epilepsy with febrile seizures [Baulac et al. (2001) Nat. Genet. 28, 46-48]. This mutation caused a decreased current response to GABA [Baulac et al. (2001) Nat. Genet. 28, 46-48]. Here we determine changes that occur in the mechanism of opening and closing of transmembrane channels formed by the GABA(A) receptor as a result of this mutation. The K289M mutation was introduced into the gamma2L subunit of the rat GABA(A) receptor, and the mutated subunit was coexpressed with the alpha1 and beta2 subunits in HEK293 cells. Transient kinetic techniques suitable for investigating reactions on cell surfaces with a microsecond-to-millisecond time resolution [Hess, G. P., and Grewer, C. (1998) Methods Enzymol. 291, 443-473] were used. They allow one to determine not only the channel-opening probability and rates of receptor desensitization but also the opening and closing rates of the mutated GABA(A) receptor channel. The channel-opening equilibrium constant of the mutated receptor was found to be 5-fold lower than that of the wild type. We calculated that this decrease in the channel-opening equilibrium accounts for the dysfunction of the mutated receptor. We discuss how a knowledge of the mechanism of the mutated receptor indicates an approach for alleviating this dysfunction.
Ramakrishnan
Latha
L
Department of Molecular Biology and Genetics, 216 Biotechnology Building, Cornell University, Ithaca, New York 14853-2703, USA.
Hess
George P
GP
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D004594:Electrophysiology; D004827:Epilepsy / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009154:Mutation / Q000235:genetics*; D010782:Photolysis; D051381:Rats; D011963:Receptors, GABA-A / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016415:Sequence Alignment
D000596:Amino Acids; D011963:Receptors, GABA-A