Histidine-834 of human erythrocyte band 3 has an essential role in the conformational changes that occur during the band 3-mediated anion exchange.
We have shown that diethyl pyrocarbonate (DEPC) inhibits band 3-mediated anion exchange and that the inhibition occurs only when histidine residue(s) is (are) modified with DEPC from the cytosolic surface of resealed ghosts [Izuhara et al. (1989) Biochemistry 28, 4725-4728]. In the present study, we have identified the DEPC-modified histidine residue as His834 using liquid chromatography with electrospray ionization mass spectrometry (LC/ESI-MS). This mild, rapid, sensitive, and quantitative method was successfully applied to analysis of the unstable DEPC-histidine adduct. The DEPC modification of His834 was pH dependent and 4,4'-dinitrostilbene-2,2'-disulfonic acid (DNDS) sensitive as previously shown. After DEPC modification, band 3-mediated anion exchange is inhibited. Consistent with previous results, we confirmed that His834 was located on the cytosolic side of the membrane and the DEPC modification of His834 had allosteric effects on the extracellular DNDS-binding site of band 3. Therefore, we conclude that His834 is located at the cytosolic surface of band 3 and is an essential residue for band 3-mediated anion exchange. We will discuss important roles of the region from TM12 to TM14 in the conformational changes that occur during the band 3-mediated anion exchange.
Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan.
Xiu Ri
XR
Jin
Yoshito
Y
Abe
Chun Yan
CY
Li
Naotaka
N
Hamasaki
D001457:Anion Exchange Protein 1, Erythrocyte / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D000838:Anions; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D004047:Diethyl Pyrocarbonate / Q000737:chemistry* / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000737:chemistry / Q000378:metabolism; D006639:Histidine / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D017136:Ion Transport; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping; D011487:Protein Conformation; D021241:Spectrometry, Mass, Electrospray Ionization; D013267:Stilbenes / Q000737:chemistry; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000737:chemistry
D001457:Anion Exchange Protein 1, Erythrocyte; D000838:Anions; D010446:Peptide Fragments; D013267:Stilbenes; D006639:Histidine; C022306:4,4'-dinitro-2,2'-stilbenedisulfonic acid; D014357:Trypsin; D004047:Diethyl Pyrocarbonate