Effects of mutations in Aedes aegypti sterol carrier protein-2 on the biological function of the protein

Biochemistry. 2010 Sep 7;49(35):7532-41. doi: 10.1021/bi902026v.

Abstract

Sterol carrier protein-2 (SCP-2) is a nonspecific intracellular lipid carrier protein. However, the molecular mechanism of ligand selectivity and the in vivo function of SCP-2 remain unclear. In this study, we used site-directed mutagenesis to investigate the ligand selectivity and in vivo function of the yellow fever mosquito sterol carrier protein-2 protein (AeSCP-2). Mutations to amino acids in AeSCP-2 known to interact with bound ligand also weakened NBD-cholesterol binding. Substitution of amino acids in the ligand cavity changed the ligand specificity of mutant AeSCP-2. Overexpressing wild-type AeSCP-2 in the Aedes aegypti cultured Aag-2 cells resulted in an increase in the level of incorporation of [(3)H]cholesterol. However, overexpressing mutants that were deleterious to the binding of NBD-cholesterol in AeSCP-2 showed a loss of ability to enhance uptake of [(3)H]cholesterol in cultured cells. Interestingly, when [(3)H]palmitic acid was used as the substrate for incorporation in vivo, there was no change in the levels of incorporation with overexpression of wild-type protein or mutated AeSCP-2s. The in vivo data suggest that AeSCP-2 is involved in sterol uptake, but not fatty acid uptake. This is the first report that the cholesterol binding ability may directly correlate with AeSCP-2's in vivo function in aiding the uptake of cholesterol.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Aedes / metabolism*
  • Animals
  • Biological Transport
  • Carrier Proteins / chemistry*
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism
  • Cholesterol / metabolism
  • Kinetics
  • Ligands
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Mutation*

Substances

  • Carrier Proteins
  • Ligands
  • sterol carrier proteins
  • Cholesterol