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Author ORCID Identifier
AccessType
Campus-Only Access for One (1) Year
Document Type
dissertation
Degree Name
Doctor of Philosophy (PhD)
Degree Program
Molecular and Cellular Biology
Year Degree Awarded
2022
Month Degree Awarded
May
First Advisor
Scott C. Garman
Subject Categories
Biochemistry, Biophysics, and Structural Biology
Abstract
Saposins are helix bundle proteins which solubilize sphingolipids and present
them to lysosomal hydrolases for catabolism. Saposin B (SapB) is an activator of
globotriaosylceramide (Gb3) catabolism by α-galactosidase A (GLA). The
mechanism by which SapB activates GLA is unknown. SapB forms dimeric
water-soluble lipoprotein complexes in vitro and presents a restrictive
conformation in crystal structures. To uncover the molecular mechanism of SapB
presenting to GLA, we subjected the fluorescent substrate derivate Gb3NBD to a
series of assays involving SapB. Firstly, we showed that SapB stably binds
Gb3NBD and presents it to GLA for cleavage in vitro. Secondly, we crystallized
SapB in the presence of Gb3NBD. Thirdly, we showed that transient interactions
between SapB and GLA can be chemically cross-linked. Fourthly, we crystallized
SapB in the presence of detergent, which detergent also led to the capture of a
binary complex between SapB and GLA. These findings provide direction for
future biochemical and structural studies on the remaining saposins and their
cognate hydrolases.
DOI
https://doi.org/10.7275/28612792
Recommended Citation
Sawyer, Thomas K., "Human Saposin B Ligand Binding and Presentation to α-Galactosidase A" (2022). Doctoral Dissertations. 2569.
https://doi.org/10.7275/28612792
https://scholarworks.umass.edu/dissertations_2/2569
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This work is licensed under a Creative Commons Attribution 4.0 License.