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Agouti signaling protein (ASP) is a secreted paracrine protein that has been widely reported to function in melanogenesis and obesity and could potentially be a core protein that regulates the color and fatty phenotype of P. sinensis. In this study, we screened out interacting proteins of ASP by combined co-immunoprecipitation mass spectrometry (CoIP-MS), yeast two hybrid (Y2H) analysis, and computational predictions. We performed docking of ASP with its well-known receptor melanocortin receptor 4 (MC4R) to predict the binding capacity and to screen out actual ASP interacting proteins, CoIP-MS was performed where identified 32 proteins that could bind with ASP and Y2H confirmed seven proteins binding with ASP directly. CoIP-MS and Y2H screening results including PPI prediction revealed that vitronectin (VTN), apolipoprotein A1 (APOA1), apolipoprotein B (APOB), and filamin B (FLNB) were the key interacting proteins of ASP. VTN, APOA1, and APOB are functional proteins in lipid metabolism and various skin disorders, suggesting ASP may function in lipid metabolism through these partners. This study provided protein-protein interaction information of ASP, and the results will promote further research into the diverse roles of ASP, as well as its binding partners, and their function in different strains of P. sinensis.
This article was published in the following journal.
Name: International journal of biological macromolecules
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A secreted protein of approximately 131 amino acids (depending on species) that regulates the synthesis of eumelanin (brown/black) pigments in MELANOCYTES. Agouti protein antagonizes the signaling of MELANOCORTIN RECEPTORS and has wide distribution including ADIPOSE TISSUE; GONADS; and HEART. Its overexpression in agouti mice results in uniform yellow coat color, OBESITY, and metabolic defects similar to type II diabetes in humans.
A secreted protein of approximately 131 amino acids that is related to AGOUTI SIGNALING PROTEIN and is also an antagonist of MELANOCORTIN RECEPTOR activity. It is expressed primarily in the HYPOTHALAMUS and the ADRENAL GLAND. As a paracrine signaling molecule, AGRP is known to regulate food intake and body weight. Elevated AGRP has been associated with OBESITY.
Large collections of small molecules (molecular weight about 600 or less), of similar or diverse nature which are used for high-throughput screening analysis of the gene function, protein interaction, cellular processing, biochemical pathways, or other chemical interactions.
Protein interaction domains of about 70-90 amino acid residues, named after a common structure found in PSD-95, Discs Large, and Zona Occludens 1 proteins. PDZ domains are involved in the recruitment and interaction of proteins, and aid the formation of protein scaffolds and signaling networks. This is achieved by sequence-specific binding between a PDZ domain in one protein and a PDZ motif in another protein.
Screening techniques first developed in yeast to identify genes encoding interacting proteins. Variations are used to evaluate interplay between proteins and other molecules. Two-hybrid techniques refer to analysis for protein-protein interactions, one-hybrid for DNA-protein interactions, three-hybrid interactions for RNA-protein interactions or ligand-based interactions. Reverse n-hybrid techniques refer to analysis for mutations or other small molecules that dissociate known interactions.
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