ABCMdb

PMID: 11248239

Roerig P, Mayerhofer P, Holzinger A, Gartner J
Characterization and functional analysis of the nucleotide binding fold in human peroxisomal ATP binding cassette transporters.
FEBS Lett. 2001 Mar 9;492(1-2):66-72., [PubMed]

Sentences

No. Mutations Sentence Comment

5 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg Mutations in conserved residues of the nucleotidases (PMP70: G478R, S572I; ALDP: G512S, S606L) altered ATPase activity. Login to comment 24 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg For the mutant constructs we selected X-ALD patient mutations in highly conserved residues in the Walker A and 19-mer region of the NBF of ALDP (G512S and S606L) and the corresponding PMP70 mutations (G478R and S572I). Login to comment 84 ABCD1 p.Gly512Ser ABCD3 p.Gly478Arg We mutated the central glycine in the Walker A motif of PMP70 and ALDP making the evolutionary severe substitutions G478R and G512S. Login to comment 85 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg Additionally, we changed the conserved serine in the 19-mer motif of PMP70 and ALDP to isoleucine (S572I) and leucine (S606L), respectively. Login to comment 86 ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile Additionally, we changed the conserved serine in the 19-mer motif of PMP70 and ALDP to isoleucine (S572I) and leucine (S606L), respectively. Login to comment 88 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg The K-subunit of L-galactosidase (L-Protein Synthetic oligonucleotide Mutation ALDP 5P-CCCCAATGGCTGCAGCAAGAGCTCCC-3P G512S 5P-GGATCCGGACAGGGAGCTCTTGCTGCAGC-3P ALDP 5P-ACTGGAAGGACGTCCTGTTGGG-3P S606L 5P-CGCCACCCAACAGGACGTCCTTCC-3P PMP70 5P-GGCTGCAGAAAGAGTTCACTTTTCCG-3P G478R 5P-GGCCATAATTCACCAAGAACACGGAAA AGTGAACTCTTTCTG-3P PMP70 5P-GACGTACTCATTGGTGGAG-3P S572I 5P-CCACCAATGAGTACGTCCATCCAATCC-3P gal) in fusion with the MBP was used as a control. Login to comment 89 ABCD1 p.Gly512Ser ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg ABCD3 p.Gly478Arg The K-subunit of L-galactosidase (LProtein Synthetic oligonucleotide Mutation ALDP 5P-CCCCAATGGCTGCAGCAAGAGCTCCC-3P G512S 5P-GGATCCGGACAGGGAGCTCTTGCTGCAGC-3P ALDP 5P-ACTGGAAGGACGTCCTGTTGGG-3P S606L 5P-CGCCACCCAACAGGACGTCCTTCC-3P PMP70 5P-GGCTGCAGAAAGAGTTCACTTTTCCG-3P G478R 5P-GGCCATAATTCACCAAGAACACGGAAA AGTGAACTCTTTCTG-3P PMP70 5P-GACGTACTCATTGGTGGAG-3P S572I 5P-CCACCAATGAGTACGTCCATCCAATCC-3P gal) in fusion with the MBP was used as a control. Login to comment 117 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu G512S and S606L cause X-ALD. Login to comment 118 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu G512S and S606L cause X-ALD. Login to comment 136 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg The S606L and G478R mutants have a decreased ATP binding a¤nity while the G512S and S572I mutants decrease the maximum velocity of ATPase activity. Login to comment 137 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg The S606L and G478R mutants have a decreased ATP binding a&#a4;nity while the G512S and S572I mutants decrease the maximum velocity of ATPase activity. Login to comment 141 ABCD1 p.Gly512Ser When testing the NBF of wild type ALDP and wild type PMP70 as well as the NBF of the mutant ALDP (G512S) against full length ALDP or full length PMP70, only background levels were detected (data not shown). Login to comment 142 ABCD1 p.Gly512Ser When testing the NBF of wild type ALDP and wild type PMP70 as well as the NBF of the mutant ALDP (G512S) against full length ALDP or full length PMP70, only background levels were detected (data not shown). Login to comment 169 ABCD1 p.Gly512Ser The missense mutation G512S in the ALD gene causes X-ALD and reduces considerable ATPase activity in our recombinant polypeptide models. Login to comment 170 ABCD1 p.Gly512Ser The missense mutation G512S in the ALD gene causes X-ALD and reduces considerable ATPase activity in our recombinant polypeptide models. Login to comment 171 ABCD1 p.Ser606Leu In contrast, the X-ALD patient mutation S606L alters ATP binding a¤nity but has no e¡ect on the overall ATPase activity. Login to comment 172 ABCD1 p.Ser606Leu ABCD1 p.Ser606Leu Because the carboxyl-terminal halves of ALDP, PMP70 and ALDR are involved in protein dimerization and interactions with other protein motifs like the transmembrane domain [28], it is possible that the S606L missense mutation in ALDP exerts its disease causing e¡ect in this manner. Login to comment 173 ABCD1 p.Ser606Leu Because the carboxyl-terminal halves of ALDP, PMP70 and ALDR are involved in protein dimerization and interactions with other protein motifs like the transmembrane domain [28], it is possible that the S606L missense mutation in ALDP exerts its disease causing e&#a1;ect in this manner. Login to comment 182 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg Previous studies on MDR suggest that the ATPase activity of the native protein Table 1 Kinetic parameters of ATPase activity in wild type and mutant ALDP and PMP70 NBF fusion proteins Fusion protein KM (WM) Vmax (nmol/Wmol NBF/min) Speci'c activity (1033 U/mg) ALDP (wild type) 11.5 þ 0.97 641.9 þ 10.7 10.0 þ 0.17 ALDP (G512S) 17.9 þ 1.23 279.3 þ 4.2 4.4 þ 0.06 ALDP (S606L) 45.6 þ 2.40 666.0 þ 8.7 10.4 þ 0.14 PMP70 (wild type) 8.2 þ 0.52 580.8 þ 6.7 9.0 þ 0.10 PMP70 (G478R) 161.8 þ 34.40 641.2 þ 28.2 10.0 þ 0.44 PMP70 (S572I) 9.9 þ 0.82 298.1 þ 4.7 4.6 þ 0.07 The kinetic data of all fusion proteins are mean values and standard deviations of 15^20 measurements at various protein concentrations using at least two distinct protein preparations. Login to comment 183 ABCD1 p.Gly512Ser ABCD1 p.Ser606Leu ABCD3 p.Ser572Ile ABCD3 p.Gly478Arg Previous studies on MDR suggest that the ATPase activity of the native protein Table 1 Kinetic parameters of ATPase activity in wild type and mutant ALDP and PMP70 NBF fusion proteins Fusion protein KM (WM) Vmax (nmol/Wmol NBF/min) Speci'c activity (1033 U/mg) ALDP (wild type) 11.5 &#fe; 0.97 641.9 &#fe; 10.7 10.0 &#fe; 0.17 ALDP (G512S) 17.9 &#fe; 1.23 279.3 &#fe; 4.2 4.4 &#fe; 0.06 ALDP (S606L) 45.6 &#fe; 2.40 666.0 &#fe; 8.7 10.4 &#fe; 0.14 PMP70 (wild type) 8.2 &#fe; 0.52 580.8 &#fe; 6.7 9.0 &#fe; 0.10 PMP70 (G478R) 161.8 &#fe; 34.40 641.2 &#fe; 28.2 10.0 &#fe; 0.44 PMP70 (S572I) 9.9 &#fe; 0.82 298.1 &#fe; 4.7 4.6 &#fe; 0.07 The kinetic data of all fusion proteins are mean values and standard deviations of 15^20 measurements at various protein concentrations using at least two distinct protein preparations. Login to comment