BACKGROUND: ABCA2 is a member of the ATP binding cassette transporter family with functional roles in cholesterol homeostasis and drug resistance. MATERIALS AND METHODS: In order to characterize its ATPase activity, we transfected HEK293 cells with an ABCA2 mammalian expression system and isolated ABCA2-enriched membranes. RESULTS: We found no measurable ATPase activity of ABCA2 in isolated membranes, except in the presence of the methyl-beta-cyclodextrin. However, competitive binding of a pseudo-substrate, 8-azido-[alpha-32P]-ATP, was demonstrated. CHO cells transfected with ABCA2 did not have a higher rate of endogenous ATP hydrolysis when compared to the mock-transfected cells. CONCLUSION: Overall, we conclude that, while ABCA2 may have low levels of ATPase activity that can be substrate-stimulated, it is more likely to have a regulatory role in cell physiology.
BACKGROUND:ABCA2 is a member of the ATP binding cassette transporter family with functional roles in cholesterol homeostasis and drug resistance. MATERIALS AND METHODS: In order to characterize its ATPase activity, we transfected HEK293 cells with an ABCA2mammalian expression system and isolated ABCA2-enriched membranes. RESULTS: We found no measurable ATPase activity of ABCA2 in isolated membranes, except in the presence of the methyl-beta-cyclodextrin. However, competitive binding of a pseudo-substrate, 8-azido-[alpha-32P]-ATP, was demonstrated. CHO cells transfected with ABCA2 did not have a higher rate of endogenous ATP hydrolysis when compared to the mock-transfected cells. CONCLUSION: Overall, we conclude that, while ABCA2 may have low levels of ATPase activity that can be substrate-stimulated, it is more likely to have a regulatory role in cell physiology.
Authors: G Szakács; T Langmann; C Ozvegy; E Orsó; G Schmitz; A Váradi; B Sarkadi Journal: Biochem Biophys Res Commun Date: 2001-11-16 Impact factor: 3.575