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Exoplasmic cysteine Cys384 of the HDL receptor SR-BI is critical for its sensitivity to a small-molecule inhibitor and normal lipid transport activity.

Publication TypeJournal Article
AuthorsYu, M., Romer KA, Nieland TJ, Xu S., Saenz-Vash V., Penman M., Yesilaltay A., Carr SA, and Krieger M.
AbstractThe HDL receptor, scavenger receptor, class B, type I (SR-BI), is a homooligomeric cell surface glycoprotein that controls HDL structure and metabolism by mediating the cellular selective uptake of lipids, mainly cholesteryl esters, from HDL. The mechanism underlying SR-BI-mediated lipid transfer, which differs from classic receptor-mediated endocytosis, involves a two-step process (binding followed by lipid transport) that is poorly understood. Our previous structure/activity analysis of the small-molecule inhibitor blocker of lipid transport 1 (BLT-1), which potently (IC(50) ∼ 50 nM) blocks SR-BI-mediated lipid transport, established that the sulfur in BLT-1's thiosemicarbazone moiety was essential for activity. Here we show that BLT-1 is an irreversible inhibitor of SR-BI, raising the possibility that cysteine(s) in SR-BI interact with BLT-1. Mass spectrometric analysis of purified SR-BI showed two of its six exoplasmic cysteines have free thiol groups (Cys251 and Cys384). Converting Cys384 (but not Cys251) to serine resulted in complete BLT-1 insensitivity, establishing that the unique molecular target of BLT-1 inhibition of cellular SR-BI dependent lipid transport is SR-BI itself. The C384S substitution reduced the receptor's intrinsic lipid uptake activity by approximately 60% without dramatically altering its surface expression, homooligomerization, or HDL binding. Thus, a small-molecule screening approach identified a key residue in SR-BI involved in lipid transport, providing a powerful springboard into the analyses of the structure and mechanism of SR-BI, and highlighting the power of this approach for such analyses.
Year of Publication2011
JournalProceedings of the National Academy of Sciences of the United States of America
Volume108
Issue30
Pages12243-8
Date Published (YYYY/MM/DD)2011/07/26
ISSN Number0027-8424
DOI10.1073/pnas.1109078108
PubMedhttp://www.ncbi.nlm.nih.gov/pubmed/21746906?dopt=Abstract