The liver isoform of carnitine palmitoyltransferase 1 is not targeted to the endoplasmic reticulum
The liver isoform of carnitine palmitoyltransferase 1 is not targeted to the endoplasmic reticulum
复制标题
DOI：
10.1042/bj20021269
复制
发表时间：
2003-02-15
影响因子：
4.1
通讯作者：
Saggerson, ED
中科院分区：
文献类型：
--
作者：
Broadway, NM;Pease, RJ;Saggerson, ED
肝微粒体部分含有丙二酰辅酶A可抑制的肉毒碱酰基转移酶（CAT）活性。已提出[Fraser、Corstorphine、Price 和 Zammit (1999) FEBS Lett。 [446, 69-74] 这种微粒体 CAT 活性是由于肝脏形式的肉毒碱棕榈酰转移酶 1 (L-CPT(1)) 可能通过 N 端信号序列靶向内质网 (ER) 膜和线粒体 [Cohen, Guillerault, Girard 和 Prip-Buus (2001) J. Biol. 446, 69-74]化学。 276, 5403-5411]。 COS-I细胞被瞬时转染以表达融合蛋白，其中增强型绿色荧光蛋白融合至L-CPT(1)的C末端。共聚焦显微镜显示该融合蛋白定位于线粒体，并且可能定位于过氧化物酶体，但不定位于内质网。对应于截短的（氨基酸 1-328）或全长 L-CPT(1) 的 cDNA 在犬胰腺微粒体存在下进行转录和翻译。然而，没有证据表明 CPT(1) 确实插入 ER 膜中。通过蔗糖密度梯度离心纯化的大鼠肝微粒体级分含有所有 88 kDa 蛋白 (p88)，该蛋白可被抗 L-CPT(1) 抗体和 2,4-二硝基苯酚-etomoxiryl-CoA（L-CPT(1) 的共价抑制剂）识别。 p88 和丙二酰辅酶 A 抑制性 CAT 活性的丰度增加了约 10%。饥饿24小时3倍。脱氧胆酸以大致相同的程度溶解了微粒体中 p88 和丙二酰辅酶 A 抑制的 CAT 活性。微粒体级分含有孔蛋白，相对于总蛋白，孔蛋白与粗制线粒体外膜级分一样丰富。结论是，L-CPT(1) 不是针对 ER 膜的，并且微粒体级分中的丙二酰辅酶 A CAT 是源自线粒体的 L-CPT(1)，可能来自膜接触位点。
Liver microsomal fractions contain a malonyl-CoA-inhibitable carnitine acyltransferase (CAT) activity. It has been proposed [Fraser, Corstorphine, Price and Zammit (1999) FEBS Lett. 446, 69-74] that this microsomal CAT activity is due to the liver form of carnitine palmitoyltransferase 1 (L-CPT(1)) being targeted to the endoplasmic reticulum (ER) membrane as well as to mitochondria, possibly by an N-terminal signal sequence [Cohen, Guillerault, Girard and Prip-Buus (2001) J. Biol. Chem. 276, 5403-5411]. COS- I cells were transiently transfected to express a fusion protein in which enhanced green fluorescent protein was fused to the C-terminus of L-CPT(1). Confocal microscopy showed that this fusion protein was localized to mitochondria, and possibly to peroxisomes, but not to the ER. cDNAs corresponding to truncated (amino acids 1-328) or full-length L-CPT(1) were transcribed and translated in the presence of canine pancreatic microsomes. However, there was no evidence of authentic insertion of CPT(1) into the ER membrane. Rat liver microsomal fractions purified by sucrose-density-gradient centrifugation contained all 88 kDa protein (p88) which was recognized by an anti-L-CPT(1) antibody and by 2,4-dinitrophenol-etomoxiryl-CoA, a covalent inhibitor of L-CPT(1). Abundance of p88 and malonyl-CoA-inhibitable CAT activity were increased approx. 3-fold by starvation for 24 h. Deoxycholate solubilized p88 and malonyl-CoA-inhibitable CAT activity from microsomes to approximately the same extent. The microsomal fraction contained porin, which, relative to total protein, was Lis abundant as in crude mitochondrial outer membranes fractions. It is concluded that L-CPT(1) is not targeted to the ER membrane and that malonyl-CoA CAT in microsomal fractions is L-CPT(1) that is derived from mitochoildria, possibly from membrane contact sites.