The first step in steroidogenesis is the movement of cholesterol from the outer to inner mitochondrial membrane; this movement is facilitated by the steroidogenic acute regulatory protein (StAR). StAR has molten-globule properties at low pH and a protease-resistant N-terminal domain at pH4 and pH8 comprising residues 63–193. To explore the mechanism of action of StAR we investigated the structural properties of the bacterially expressed N-terminal domain (63–193 StAR) using CD, limited proteolysis and NMR. Far- and near-UV CD showed that the amount of secondary structure was greater at acidic than at neutral pH, but there was little tertiary structure at any pH. Unlike 63–193 StAR liberated from N-62 StAR by proteolysis, biosynthetic 63–193 StAR was no longer resistant to trypsin or proteinase K at pH7, or to pepsin at pH4. Addition of trifluoroethanol and SDS increased secondary structure at pH7, and dodecylphosphocholine and CHAPS increased secondary structure at pH2, pH4 and pH7. However, none of these conditions induced tertiary structure, as monitored by near-UV CD or NMR. Liposomes of phosphatidylcholine, phosphatidylserine and their mixture increased secondary structure of 63–193 StAR at pH7, as monitored by far-UV CD, and stable protein–liposome complexes were identified by gel-permeation chromatography. These results provide further evidence that the N-terminal domain of StAR is a molten globule, and provide evidence that this conformation facilitates the interaction of the N-terminal domain of StAR with membranes. We suggest that this interaction is the key to understanding the mechanism of StAR's action.
Molten-globule structure and membrane binding of the N-terminal protease-resistant domain (63–193) of the steroidogenic acute regulatory protein (StAR)
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Maengseok SONG, Haiyan SHAO, Anwer MUJEEB, Thomas L. JAMES, Walter L. MILLER; Molten-globule structure and membrane binding of the N-terminal protease-resistant domain (63–193) of the steroidogenic acute regulatory protein (StAR). Biochem J 15 May 2001; 356 (1): 151–158. doi: https://doi.org/10.1042/bj3560151
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