Ligand binding and structural analysis of a human putative cellular retinol-binding protein.
Three cellular retinol-binding protein (CRBP) types (CRBP I, II, and III) with distinct tissue distributions and retinoid binding properties have been structurally characterized thus far. A human binding protein, whose mRNA is expressed primarily in kidney, heart, and transverse colon, is shown here to be a CRBP family member (human CRBP IV), according to amino acid sequence, phylogenetic analysis, gene structure organization, and x-ray structural analysis. Retinol binding to CRBP IV leads to an absorption spectrum distinct from a typical holo-CRBP spectrum and is characterized by an affinity (K(d) = approximately 200 nm) lower than those for CRBP I, II, and III, as established in direct and competitive binding assays. As revealed by mutagenic analysis, the presence in CRBP IV of His(108) in place of Gln(108) is not responsible for the unusual holo-CRBP IV spectrum. The 2-A resolution crystal structure of human apo-CRBP IV is very similar to those of other structurally characterized CRBPs. The side chain of Tyr(60) is present within the binding cavity of the apoprotein and might affect the interaction with the retinol molecule. These results indicate that human CRBP IV belongs to a clearly distinct CRBP subfamily and suggest a relatively different mode of retinol binding for this binding protein.
Folli
Claudia
C
Department of Biochemistry and Molecular Biology, University of Parma, Italy.
Calderone
Vito
V
Ramazzina
Ileana
I
Zanotti
Giuseppe
G
Berni
Rodolfo
R
D000595:Amino Acid Sequence; D003460:Crystallization; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation; D012333:RNA, Messenger / Q000032:analysis; D011994:Recombinant Proteins / Q000378:metabolism; D012177:Retinol-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054840:Retinol-Binding Proteins, Cellular; D014801:Vitamin A / Q000378:metabolism
D012333:RNA, Messenger; D011994:Recombinant Proteins; D012177:Retinol-Binding Proteins; D054840:Retinol-Binding Proteins, Cellular; D014801:Vitamin A