ATP Binding/Hydrolysis by and Phosphorylation of Peroxisomal ATP-binding Cassette Proteins PMP70 (ABCD3) and Adrenoleukodystrophy Protein (ABCD1)*
ATP Binding/Hydrolysis by and Phosphorylation of Peroxisomal ATP-binding Cassette Proteins PMP70 (ABCD3) and Adrenoleukodystrophy Protein (ABCD1)*
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DOI：
10.1074/jbc.m205079200
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发表时间：
2002-10
期刊：
影响因子：
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通讯作者：
Arowu R. Tanaka;K. Tanabe;M. Morita;Mikinori Kurisu;Yoshinori Kasiwayama;M. Matsuo;N. Kioka;T. Amachi;T. Imanaka;K. Ueda
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作者：
Arowu R. Tanaka;K. Tanabe;M. Morita;Mikinori Kurisu;Yoshinori Kasiwayama;M. Matsuo;N. Kioka;T. Amachi;T. Imanaka;K. Ueda
70 kDa过氧化物酶体膜蛋白（PMP 70）和肾上腺脑白质营养不良蛋白（ALDP），一半大小的ATP结合盒转运蛋白，参与长链和超长链脂肪酸代谢转运到过氧化物酶体。我们研究的相互作用的过氧化物酶体ATP结合盒转运ATP使用大鼠肝脏过氧化物酶体。当过氧化物酶体与8-叠氮基-[α-32 P]ATP和8-叠氮基-[γ-32 P]ATP在37 °C下在不存在Mg 2+的情况下孵育并暴露于UV光而不去除未结合的核苷酸时，PMP 70以相似的效率被光亲和标记。光亲和标记的PMP 70和ALDP与其他过氧化物酶体蛋白一起进行免疫共沉淀，这些蛋白也显示出紧密的ATP结合特性。Mg ~（2+）使PMP 70与8-叠氮基-[γ-32 P]ATP的光亲和标记减少70%，而与8-叠氮基-[α-32 P]ATP的光亲和标记仅减少20%。然而，三分之二的核苷酸（可能是ADP）在未结合的核苷酸的去除过程中解离。这些结果表明，ATP与PMP 70紧密结合，在Mg 2+的存在下，结合的ATP被水解为ADP，并且产生的ADP从PMP 70解离，这允许ATP水解周转。ALDP的光亲和标记特性与PMP 70基本相似。在PMP 70和ALDP中未观察到钒酸盐诱导的核苷酸捕获。PMP 70和ALDP也在酪氨酸残基处磷酸化。PMP 70和ALDP的ATP结合/水解和磷酸化参与脂肪酸转运到过氧化物酶体的调节。
The 70-kDa peroxisomal membrane protein (PMP70) and adrenoleukodystrophy protein (ALDP), half-size ATP-binding cassette transporters, are involved in metabolic transport of long and very long chain fatty acids into peroxisomes. We examined the interaction of peroxisomal ATP-binding cassette transporters with ATP using rat liver peroxisomes. PMP70 was photoaffinity-labeled at similar efficiencies with 8-azido-[α-32P]ATP and 8-azido-[γ-32P]ATP when peroxisomes were incubated with these nucleotides at 37 °C in the absence Mg2+ and exposed to UV light without removing unbound nucleotides. The photoaffinity-labeled PMP70 and ALDP were co-immunoprecipitated together with other peroxisomal proteins, which also showed tight ATP binding properties. Addition of Mg2+ reduced the photoaffinity labeling of PMP70 with 8-azido-[γ-32P]ATP by 70%, whereas it reduced photoaffinity labeling with 8-azido-[α-32P]ATP by only 20%. However, two-thirds of nucleotide (probably ADP) was dissociated during removal of unbound nucleotides. These results suggest that ATP binds to PMP70 tightly in the absence of Mg2+, the bound ATP is hydrolyzed to ADP in the presence of Mg2+, and the produced ADP is dissociated from PMP70, which allows ATP hydrolysis turnover. Properties of photoaffinity labeling of ALDP were essentially similar to those of PMP70. Vanadate-induced nucleotide trapping in PMP70 and ALDP was not observed. PMP70 and ALDP were also phosphorylated at a tyrosine residue(s). ATP binding/hydrolysis by and phosphorylation of PMP70 and ALDP are involved in the regulation of fatty acid transport into peroxisomes.