ABCMdb

PMID: 22735453

Quazi F, Lenevich S, Molday RS
ABCA4 is an N-retinylidene-phosphatidylethanolamine and phosphatidylethanolamine importer.
Nat Commun. 2012 Jun 26;3:925. doi: 10.1038/ncomms1927., [PubMed]

Sentences

No. Mutations Sentence Comment

66 ABCA4 p.Lys1978Met ABCA4 p.Lys969Met This was examined using donor proteoliposomes reconstituted with wild-type (WT) ABCA4 or the ATPase deficient K969M/K1978M double mutant (ABCA4-MM) (Fig. 2d). Login to comment 93 ABCA4 p.Lys1978Met ABCA4 p.Lys969Met (d) Effect of ATP, ADP and AMP-PNP on [3H] ATR transfer from donor proteoliposomes reconstituted with WT ABCA4 or the ATPase-impaired K969M/K1978M double mutant (ABCA4-MM) purified from transfected HEK293 cells. Login to comment 145 ABCA4 p.Gly863Ala ABCA4 p.Asn965Ser ABCA4 p.Lys1978Met ABCA4 p.Lys969Met ABCA4 p.Lys969Met To gain further insight into the molecular mechanisms underlying ABCA4-mediated transport activity and Stargardt disease, we examined the functional properties of ABCA4-containing G863A and N965S mutations associated with Stargardt disease and Walker A mutations, K969M in NBD1, K1978M in NBD2 and the double mutant K969M/1978M. Login to comment 146 ABCA4 p.Gly863Ala ABCA4 p.Asn965Ser ABCA4 p.Lys1978Met ABCA4 p.Lys1978Met ABCA4 p.Lys969Met ABCA4 p.Lys969Met The K1978M mutant expressed at the same level as WT ABCA4, whereas the G863A, N965S, K969M and K969M/ K1978M mutants expressed within 50% that of WT ABCA4. Login to comment 168 ABCA4 p.Gly863Ala ABCA4 p.Gly863Ala ABCA4 p.Asn965Ser ABCA4 p.Asn965Ser Percent-relative NBD-PE flipping plotted as a mean with error bars representing ± s.d. for n = 3. mutants showing essentially undetectable activity and the G863A and N965S Stargardt mutants displaying ~18 and 37% of WT activity (Fig. 6b). Login to comment 172 ABCA4 p.Gly863Ala ABCA4 p.Asn965Ser ABCA4 p.Lys1978Met ABCA4 p.Lys1978Met ABCA4 p.Lys969Met ABCA4 p.Lys969Met Addition of ATP resulted in release of the substrate from the G863A, N965S and K1978M mutants, but impaired release from the K969M mutant and no significant release from the K969M/K1978M double mutant. Login to comment 189 ABCA4 p.Gly863Ala ABCA4 p.Asn965Ser ABCA4 p.Lys1978Met ABCA4 p.Lys1978Met ABCA4 p.Lys969Met ABCA4 p.Lys969Met However, retinal readily reacts with PE, I a E ABCA4 W i l d - t y p e G 8 6 3 A N 9 6 5 S K 9 6 9 M K 9 6 9 M / K 1 9 7 8 M K 1 9 7 8 M ABCA4 kDa 250 250 I E I E I E I E I E 150 100 75 50 37 25 e % N-retinylidene PE binding 120 - ATP + ATP 100 80 60 40 20 0 G 8 6 3 A M M N 9 6 5 S K 9 6 9 M K 1 9 7 8 M W T b G 8 6 3 A M M N 9 6 5 S K 9 6 9 M K 1 9 7 8 M 120 % Retinal transfer * * 100 80 60 40 20 0 W T c G863A N965S Retinal (µM) 250 % Basal ATPase activity 200 150 100 WT 50 0 0 10 20 30 40 50 60 d MM Retinal (µM) K969M K1978M % Basal ATPase activity 250 WT 200 150 100 50 0 0 10 20 30 40 50 60 f % NBD-PE flippase activity * * 120 100 80 60 40 20 0 G 8 6 3 A M M N 9 6 5 S K 9 6 9 M K 1 9 7 8 M W T Figure 6 | Effect of Walker A and Stargardt mutations on ATR transfer activity. Login to comment 219 ABCA4 p.Gly863Ala ABCA4 p.Gly863Ala ABCA4 p.Asn965Ser ABCA4 p.Asn965Ser The binding of N-retinyli- dene-PE and its release by ATP is not affected by the G863A and N965S mutations (Fig. 6e), but the basal- and retinal-stimulated ATPase activity and ATP-dependent retinal transfer activity are significantly reduced. Login to comment