Effects of amino acid substitutions at glycine 420 on SR-BI cholesterol transport function.
Scavenger receptor class B type I (SR-BI) facilitates the uptake of HDL cholesteryl esters (CEs) in a two-step process involving binding of HDL to its extracellular domain and transfer of HDL core CEs to a metabolically active membrane pool, where they are subsequently hydrolyzed by a neutral CE hydrolase. Recently, we characterized a mutant, G420H, which replaced glycine 420 in the extracellular domain of SR-BI with a histidine residue and had a profound effect on SR-BI function. The G420H mutant receptor exhibited a reduced ability to mediate selective HDL CE uptake and was unable to deliver HDL CE for hydrolysis, despite the fact that it retained the ability to bind HDL. This did not hold true if glycine 420 was replaced with an alanine residue; G420A maintained wild-type HDL binding and cholesterol transport activity. To further understand the role that glycine 420 plays in SR-BI function and why there was a disparity between replacing glycine 420 with a histidine versus an alanine, we generated a battery of point mutants by substituting glycine 420 with amino acids possessing side chains that were charged, hydrophobic, polar, or bulky and tested the resulting mutants for their ability to support HDL binding, HDL cholesterol transport, and delivery for hydrolysis. The results indicated that substitution with a negatively charged residue or a proline impaired cell surface expression of SR-BI or its interaction with HDL, respectively. Furthermore, substitution of glycine 420 with a positively charged residue reduced HDL CE uptake as well as its subsequent hydrolysis.
Parathath
Saj
S
Department of Pharmacological Sciences, University Medical Center, Stony Brook University, Stony Brook, NY 11794-8651, USA.
Darlington
Yolanda F
YF
de la Llera Moya
Margarita
M
Drazul-Schrader
Denise
D
Williams
David L
DL
Phillips
Michael C
MC
Rothblat
George H
GH
Connelly
Margery A
MA
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001692:Biological Transport / Q000235:genetics / Q000502:physiology; D001710:Biotin / Q000378:metabolism; D019556:COS Cells; D002522:Chlorocebus aethiops; D002784:Cholesterol / Q000378:metabolism; D002788:Cholesterol Esters / Q000378:metabolism / Q000493:pharmacokinetics; D005998:Glycine / Q000235:genetics*; D006639:Histidine / Q000235:genetics / Q000502:physiology; D006801:Humans; D008075:Lipoproteins, HDL / Q000378:metabolism / Q000493:pharmacokinetics; D008969:Molecular Sequence Data; D017354:Point Mutation; D051122:Scavenger Receptors, Class B / Q000235:genetics* / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D002788:Cholesterol Esters; C059714:HDL cholesteryl ester; D008075:Lipoproteins, HDL; D051122:Scavenger Receptors, Class B; D006639:Histidine; D001710:Biotin; D002784:Cholesterol; D005998:Glycine