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Identification of Galectin-3 As a High-Affinity Binding Protein for Advanced Glycation End Products (AGE): A New Member of the AGE-Receptor Complex

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Abstract

Background

Advanced glycation end products (AGE), the reactive derivatives of nonenzymatic glucose-protein condensation reactions, are implicated in the multiorgan complications of diabetes and aging. An AGE-specific cellular receptor complex (AGE-R) mediating AGE removal as well as multiple biological responses has been identified. By screening an expression library using antibody against a previously identified component of the AGE-R complex p90, a known partial cDNA clone was isolated with homology to galectin-3, a protein of diverse identity, and member of the galectin family.

Materials and Methods

To explore this unexpected finding, the nature of the interactions between galectin-3 and AGE was studied using intact macrophage-like RAW 264.7 cells, membrane-associated and recombinant galectin-1 through -4, and model AGE-ligands (AGE-BSA, FFI-BSA).

Results

Among the members of this family (galectin-1 through 4), recombinant rat galectin-3 was found to exhibit high-affinity 125I-AGE-BSA binding with saturable kinetics (kD 3.5 × 107 M−1) that was fully blocked by excess unlabeled naturally formed AGE-BSA or synthetic FFI-BSA, but only weakly inhibited by several known galectin-3 ligands, such as lactose. In addition to the p90, immunoprecipitation with anti-galectin-3, followed by 125I-AGE-BSA ligand blot analysis of RAW 264.7 cell extracts, revealed galectin-3 (28 and 32 kD), as well as galectin-3-associated proteins (40 and 50 kD) with AGE-binding activity. Interaction of galectin-3 with AGE-BSA or FFI-BSA resulted in formation of SDS-, and β-mercaptoethanol-insoluble, but hydroxylamine-sensitive high-molecular weight complexes between AGE-ligand, galectin-3, and other membrane components.

Conclusions

The findings point toward a mechanism by which galectin-3 may serve in the assembly of AGE-R components and in the efficient cell surface attachment and endocytosis by macrophages of a heterogenous pool of AGE moieties with diverse affinities, thus contributing to the elimination of these pathogenic substances.

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Acknowledgments

We would like to thank Dr. Kirk Manogue and Dr. Bradley Berger for their help and guidance. We also thank Dr. Hakon Leffler and Dr. Samuel H. Barondes (University of California, San Francisco, CA) for the supply of the galectin 1–4 and their valuable cooperation. Our thanks to Dr. J. L. Wang (Michigan State University) for providing CBP-35, and C-peptide and to Dr. Vincent Monnier (Case Western Reserve, CL) for pyrraline and pentosidine which were also generously provided. These studies were supported in part by The National Institutes of Health Grants AGO-6943 and AGO-9453 to HV.

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  1. The Picower Institute for Medical Research, 350 Community Drive, Manhasset, New York, 11030, USA

    Helen Vlassara, Yong Ming Li, Farhad Imani, Donald Wojciechowicz, Zhi Yang & Anthony Cerami

  2. The Scripps Research Institute, La Jolla, California, USA

    Fu-Tong Liu

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  1. Helen Vlassara
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Vlassara, H., Li, Y.M., Imani, F. et al. Identification of Galectin-3 As a High-Affinity Binding Protein for Advanced Glycation End Products (AGE): A New Member of the AGE-Receptor Complex. Mol Med 1, 634–646 (1995). https://doi.org/10.1007/BF03401604

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