9AXG

Human saposin B in the presence of globotriaosylceramide-NBD

Summary for 9AXG

• Entry DOI: 10.2210/pdb9axg/pdb
• Related: 9AVS
• Descriptor: Saposin-B, MALONATE ION (3 entities in total)
• Functional Keywords: saposin, saposin b, sapb, fabry disease, lysosomal, activator protein, globotriaosylceramide, gb3, nitrobenzoxadiazole, nbd, lipid binding protein
• Biological source: Homo sapiens (human)
• Total number of polymer chains: 2
• Total formula weight: 18296.95

Authors

Sawyer, T.K.,Garman, S.C. (deposition date: 2024-03-06, release date: 2024-03-13, Last modification date: 2024-11-20)

Primary citation

Sawyer, T.K.,Aral, E.,Staros, J.V.,Bobst, C.E.,Garman, S.C.
Human Saposin B Ligand Binding and Presentation to alpha-Galactosidase A.
Biorxiv, 2024

PubMed Abstract: Sphingolipid activator protein B (saposin B; SapB) is an essential activator of globotriaosylceramide (Gb3) catabolism by α-galactosidase A. However, the manner by which SapB stimulates α-galactosidase A activity remains unknown. To uncover the molecular mechanism of SapB presenting Gb3 to α-galactosidase A, we subjected the fluorescent substrate globotriaosylceramide-nitrobenzoxidazole (Gb3-NBD) to a series of biochemical and structural assays involving SapB. First, we showed that SapB stably binds Gb3-NBD using a fluorescence equilibrium binding assay, isolates Gb3-NBD from micelles, and facilitates α-galactosidase A cleavage of Gb3-NBD . Second, we crystallized SapB in the presence of Gb3-NBD and validated the ligand-bound assembly. Third, we captured transient interactions between SapB and α-galactosidase A by chemical cross-linking. Finally, we determined the crystal structure of SapB bound to α-galactosidase A. These findings establish general principles for molecular recognition in saposin:hydrolase complexes and highlight the utility of NBD reporter lipids in saposin biochemistry and structural biology.

PubMed: 38617236
DOI: 10.1101/2024.04.04.584535

Experimental method

X-RAY DIFFRACTION (2.68 Å)