title: Characterization of band 3-ankyrin-Protein 4.2 complex by biochemical and mass spectrometry approaches.
abstract: The elastic property of red blood cell is supported by interaction between red cell membrane and the intricate cytoskeleton network underlying the membrane bilayer cytoplasmic face. One of the major scaffold protein linkers is band 3-ankyrin complex. Defects occurring in this complex have been found in many inherited diseases, causing red blood cell abnormalities. Here we combined the power of mass spectrometry with conventional biochemical purification methods in order to study the native interactions among band 3, ankyrin and Protein 4.2. This approach provided in vivo evidence for the association between band 3 and N-terminal ankyrin purified directly from the cell membrane. The C-terminal regions of ankyrin were not found to be a stable partner of the band 3 complex. Protein 4.2 was shown here to be an integral part of the complex. Its association to the band 3-ankyrin complex could withstand harsh purification conditions. Our findings lend additional support to the interaction between band 3 and ankyrin N-terminal domain previously shown by in vitro binding assays and provide evidence for a band 3 core complex comprising of band 3, ankyrin and Protein 4.2.
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok 10400, Thailand.
authors.forename: Krittikorn
authors.initials: K
authors.lastname: Kümpornsin
authors.forename: Surasak
authors.initials: S
authors.lastname: Jiemsup
authors.forename: Suganya
authors.initials: S
authors.lastname: Yongkiettrakul
authors.forename: Thanat
authors.initials: T
authors.lastname: Chookajorn
mesh_terms: D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry* / Q000302:isolation & purification; D017487:Ankyrins / Q000737:chemistry* / Q000302:isolation & purification; D002853:Chromatography, Liquid; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000302:isolation & purification; D004910:Erythrocyte Membrane / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000302:isolation & purification; D017434:Protein Structure, Tertiary
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D017487:Ankyrins; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D046912:Multiprotein Complexes; C044791:erythrocyte membrane band 4.2 protein