Protein 4.1 deficiency associated with an altered binding to the spectrin-actin complex of the red cell membrane skeleton.
Protein 4.1 has been defined as a major component of the subcortical skeleton of erythrocytes. It binds the spectrin--actin scaffold through a 10-kD internal domain. This binding requires an essential 21-amino acid sequence motif, Motif I, which is retained by alternative splicing at the late stage of erythroid differentiation. We here analyze the molecular basis of heterozygous 4.1(-) hereditary elliptocytosis, associated with protein 4.1 partial deficiency, in nine related French families. cDNA sequencing revealed a single codon deletion (AAA) resulting in a lysine residue deletion within the 10-kD binding domain, 3' of Motif I. The mutated allele was designated allele 4.1 Aravis. In order to assess the functional effect of the codon deletion, recombinant 10-kD constructs were made and various binding assays were performed using spectrin, purified spectrin-actin complex, or red cell membranes. These experiments demonstrated that the deletion of the Lys residue clearly prevents the binding capacity. Similar results were obtained with a construct containing the Lys residue but lacking Motif I. These data strongly suggest that the binding site to the spectrin-actin complex must contain the Lys 447 (or 448), and therefore resides not only on Motif I but extends 3' of this essential motif.
Lorenzo
F
F
CNRS URA 1171, Institut Pasteur de Lyon, France.
Dalla Venezia
N
N
Morlé
L
L
Baklouti
F
F
Alloisio
N
N
Ducluzeau
M T
MT
Roda
L
L
Lefrançois
P
P
Delaunay
J
J
D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins; D004252:DNA Mutational Analysis; D004612:Elliptocytosis, Hereditary / Q000097:blood / Q000235:genetics*; D004910:Erythrocyte Membrane / Q000378:metabolism*; D005260:Female; D005602:France; D006801:Humans; D008239:Lysine / Q000502:physiology; D008297:Male; D008565:Membrane Proteins / Q000737:chemistry / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009479:Neuropeptides; D010375:Pedigree; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins; D017384:Sequence Deletion / Q000235:genetics; D013049:Spectrin / Q000378:metabolism*
D000199:Actins; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D009479:Neuropeptides; D011993:Recombinant Fusion Proteins; C028841:erythrocyte membrane band 4.1 protein; C120557:erythrocyte membrane protein band 4.1-like 1; D013049:Spectrin; D008239:Lysine