title: Molecular basis and functional consequences of the dominant effects of the mutant band 3 on the structure of normal band 3 in Southeast Asian ovalocytosis.
abstract: Southeast Asian ovalocytosis (SAO) human red cell membranes contain similar proportions of normal band 3 and a mutant band 3 with a nine amino acid deletion (band 3 SAO). We employed specific chemical modification and proteolytic cleavage to probe the structures of band 3 in normal and SAO membranes. When the membranes were modified specifically at lysine residues with N-hydroxysulfosuccinimide-SS-biotin, band 3 Lys-851 was not modified in normal membranes but quantitatively modified in SAO membranes. Normal and SAO membranes showed different patterns of band 3 proteolytic cleavage. Notably, many sites cleaved in normal membranes were not cleaved in SAO membranes, despite the presence of normal band 3 in these membranes. The mutant band 3 changes the structure of essentially all the normal band 3 present in the SAO membranes, and these changes extend throughout the normal band 3 molecules. The results also imply that band 3 in SAO membranes is present as hetero-tetramers or higher hetero-oligomers. The dominant structural effects of band 3 SAO on the other band 3 allele have important consequences on the functional and hematological properties of human red cells heterozygous for band 3 SAO. Analysis of the altered profile of biotinylation and protease cleavage sites suggests the location of exposed surfaces in the band 3 membrane domain and identifies likely interacting regions within the molecule. Our approach provides a sensitive method for studying structural changes in polytopic membrane proteins.
authors.affiliation: Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kuma
authors.forename: Yoshito
authors.initials: Y
authors.lastname: Abe
authors.forename: David
authors.initials: D
authors.lastname: Askin
authors.forename: Lesley J
authors.initials: LJ
authors.lastname: Bruce
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Hamasaki
authors.forename: Michael J A
authors.initials: MJ
authors.lastname: Tanner
authors.forename: Naotaka
authors.initials: N
authors.lastname: Hamasaki
mesh_terms: D000595:Amino Acid Sequence; D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001710:Biotin / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004612:Elliptocytosis, Hereditary / Q000235:genetics*; D004910:Erythrocyte Membrane / Q000378:metabolism; D004913:Erythrocytes, Abnormal / Q000378:metabolism*; D006579:Heterozygote; D006801:Humans; D006868:Hydrolysis; D008239:Lysine / Q000737:chemistry; D008969:Molecular Sequence Data; D009154:Mutation; D013388:Succinimides / Q000737:chemistry
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D013388:Succinimides; D001710:Biotin; C035761:N-hydroxysulfosuccinimide; D008239:Lysine