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Role of Surface Lysine Residues of Adipocyte Fatty Acid-Binding Protein in Fatty Acid Transfer to Phospholipid Vesicles

2001; American Chemical Society; Volume: 40; Issue: 21 Linguagem: Inglês

10.1021/bi0101042

1943-295X

Heng-Ling Liou, Judith Storch,

Adipose Tissue and Metabolism

The tertiary structure of murine adipocyte fatty acid-binding protein (AFABP) is a flattened 10-stranded β-barrel capped by a helix−turn−helix segment. This helical domain is hypothesized to behave as a "lid" or portal for ligand entry into and exit from the binding cavity. Previously, we demonstrated that anthroyloxy-labeled fatty acid (AOFA) transfer from AFABP to phospholipid membranes occurs by a collisional process, in which ionic interactions between positively charged lysine residues on the protein surface and negatively charged phospholipid headgroups are involved. In the present study, the role of specific lysine residues located in the portal and other regions of AFABP was directly examined using site-directed mutagenesis. The results showed that isoleucine replacement for lysine in the portal region, including the αI- and αII-helices and the β C-D turn, resulted in much slower 2-(9-anthroyloxy)palmitate (2AP) transfer rates to acidic membranes than those of native AFABP. An additive effect was found for mutant K22,59I, displaying the slowest rates of FA transfer. Rates of 2AP transfer from "nonportal" mutants on the β-G and I strands were affected only moderately; however, a lysine → isoleucine mutation in the nonportal β-A strand decreased the 2AP transfer rate. These studies suggest that lysines in the helical cap domain are important for governing ionic interactions between AFABP and membranes. Furthermore, it appears that more than one distinct region, including the αI-helix, αII-helix, β C-D turn, and the β-A strand, is involved in these charge−charge interactions.