biostudies-literatureByrd KMMichael, Marcia and Christa Parseghian EndowmentAra Parseghian Medical Research FoundationNational Institutes of HealthNIGMS NIH HHSUniversity of Notre Dame3843-51https://www.ebi.ac.uk/biostudies/studies/S-EPMC4461494biostudies-literatureUnknown23(13)A crosslinker was designed and synthesized as a molecular tool for potential use in probing the intracellular trafficking pathways of steroids. The design was guided by computational modeling based upon a model for the transfer of cholesterol between two proteins, NPC1 and NPC2. These proteins play critical roles in the transport of low-density lipoprotein-derived cholesterol from the lumen of lysosomes to other subcellular compartments. Two modified cholesterol residues were covalently joined by a tether based on molecular modeling of the transient interaction of NPC1 and NPC2 during the transfer of cholesterol from the binding site of one of these proteins to the other. With two cholesterol molecules appropriately connected, we hypothesize that the cholesterol binding sites of both proteins will be simultaneously occupied in a manner that will stabilize the protein-protein interaction to permit detailed structural analysis of the resulting complex. A photoaffinity label has also been introduced into one of the cholesterol cores to permit covalent attachment of one of the units into its respective protein-binding pocket. The basic design of these crosslinkers should render them useful for examining interactions of the NPC1/NPC2 pair as well as other sterol transport proteins.Bioorganic & medicinal chemistryDesign and synthesis of a crosslinker for studying intracellular steroid trafficking pathways.PMC4461494T32 GM075762T32GM075762Byrd KMWiest OEstiu GArieno MDKennelly MEHelquist PfalseDesign and synthesis of a crosslinker for studying intracellular steroid trafficking pathways.A crosslinker was designed and synthesized as a molecular tool for potential use in probing the intracellular trafficking pathways of steroids. The design was guided by computational modeling based upon a model for the transfer of cholesterol between two proteins, NPC1 and NPC2. These proteins play critical roles in the transport of low-density lipoprotein-derived cholesterol from the lumen of lysosomes to other subcellular compartments. Two modified cholesterol residues were covalently joined by a tether based on molecular modeling of the transient interaction of NPC1 and NPC2 during the transfer of cholesterol from the binding site of one of these proteins to the other. With two cholesterol molecules appropriately connected, we hypothesize that the cholesterol binding sites of both proteins will be simultaneously occupied in a manner that will stabilize the protein-protein interaction to permit detailed structural analysis of the resulting complex. A photoaffinity label has also been introduced into one of the cholesterol cores to permit covalent attachment of one of the units into its respective protein-binding pocket. The basic design of these crosslinkers should render them useful for examining interactions of the NPC1/NPC2 pair as well as other sterol transport proteins.2015-01-01T00:00:00Z2015 Jul2024-10-14T14:44:13.985Z2019-03-27T01:53:10ZS-EPMC44614942589069610.1016/j.bmc.2015.03.053