Two different members of the fatty acid‐binding protein (FABP) family are found in enterocyte cells of the gastrointestinal system, namely liver‐type and intestinal fatty acid‐binding proteins (LFABP and IFABP, also called FABP1 and FABP2, respectively). Striking phenotypic differences have been observed in knockout mice for either protein, for example, high fat‐fed IFABP‐null mice remained lean, whereas LFABP‐null mice were obese, correlating with differences in food intake. This finding prompted us to investigate the role each protein plays in directing the specificity of binding to ligands involved in appetite regulation, such as fatty acid ethanolamides and related endocannabinoids. We determined the binding affinities for nine structurally related ligands using a fluorescence competition assay, revealing tighter binding to IFABP than LFABP for all ligands tested. We found that the head group of the ligand had more impact on binding affinity than the alkyl chain, with the strongest binding observed for the carboxyl group, followed by the amide, and then the glycerol ester. These trends were confirmed using two‐dimensional ¹H–¹⁵N nuclear magnetic resonance (NMR) to monitor chemical shift perturbation of the protein backbone resonances upon titration with ligand. Interestingly, the NMR data revealed that different residues of IFABP were involved in the coordination of endocannabinoids than those implicated for fatty acids, whereas the same residues of LFABP were involved for both classes of ligand. In addition, we identified residues that are uniquely affected by binding of all types of ligand to IFABP, suggesting a rationale for its tighter binding affinity compared with LFABP.

中文翻译：

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肠道中表达的两种脂肪酸结合蛋白与内源性大麻素的相互作用不同。

在胃肠系统的肠上皮细胞中发现了脂肪酸结合蛋白（FABP）家族的两个不同成员，即肝型和肠道脂肪酸结合蛋白（LFABP和IFABP，分别称为FABP1和FABP2）。在敲除小鼠中观察到两种蛋白质的惊人表型差异，例如，高脂喂养的IFABP无效小鼠保持瘦，而LFABP无效小鼠肥胖，这与食物摄入的差异有关。这一发现促使我们研究每种蛋白质在指导与食欲调节相关配体（例如脂肪酸乙醇酰胺和相关内源性大麻素）的结合特异性中所起的作用。我们使用荧光竞争测定法确定了九种结构相关配体的结合亲和力，对于所有测试的配体来说，与LFABP的结合更紧密。我们发现配体的头基对结合亲和力的影响比烷基链更大，其中羧基的结合力最强，其次是酰胺，然后是甘油酯。这些趋势已通过二维证实¹ H– ¹⁵ N核磁共振（NMR），用于监测配体滴定后蛋白质骨架共振的化学位移扰动。有趣的是，NMR数据显示，与脂肪酸相关的IFABP残基参与的内环大麻素配位作用不同，而两种配体均涉及相同的LFABP残基。此外，我们鉴定出了受所有类型配体与IFABP结合唯一影响的残基，这表明其与LFABP相比具有更严格的结合亲和力。