Mutation of His 834 in human anion exchanger 1 affects substrate binding.
Anion exchanger 1 (AE1 or band 3) is responsible for Cl(-)-HCO3(-) exchange on erythrocyte membrane. Previously, we showed that band 3 is fixed in an inward-facing conformation by specific modification of His 834 with DEPC, resulting in a strong inhibition of its anion transport activity. To clarify the physiological role of His 834, we evaluated the sulfate transport activities of various band 3 mutants: different mutants at His 834 and alanine mutants of peripheral residues around 834 (Lys 829-Phe 836) in yeast cell membranes. The K(m) values of the His 834 mutants were 4-10 times higher than that of the wild type, while their V(max) values were barely lower than that of wild type. Meanwhile, the K(m) values of the peripheral alanine mutants were only slightly increased. These data suggest that His 834 is critically important for the efficient binding of sulfate anion, but not for the conformational change induced by substrate binding.
Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan.
Shinya
S
Takazaki
Yoshito
Y
Abe
Tomohiro
T
Yamaguchi
Mikako
M
Yagi
Tadashi
T
Ueda
Dongchon
D
Kang
Naotaka
N
Hamasaki
D000595:Amino Acid Sequence; D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006639:Histidine / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D013431:Sulfates / Q000378:metabolism
D001457:Anion Exchange Protein 1, Erythrocyte; D013431:Sulfates; D006639:Histidine