Mendoza J A, Grant E, Horowitz P M
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78284-7760.
J Protein Chem. 1993 Feb;12(1):65-9. doi: 10.1007/BF01024916.
Rhodanese (thiosulfate cyanide sulfurtransferase; E.C. 2.8.1.1) is a mitochondrial enzyme that is unprocessed after import. We describe in vitro experiments showing that partially folded rhodanese can interact with lipid bilayers. The interaction was monitored by measuring the ability of rhodanese to disrupt small unilamellar vesicles composed of phosphatidylserine and to release 6-carboxyfluorescein that was trapped in the liposomes. Partially folded rhodanese, derived by dilution of urea-unfolded enzyme, efficiently induced liposome leakage. Native rhodanese had no effect on liposome integrity. Liposome disruption progressively decreased as rhodanese was given the opportunity to refold or aggregate before introduction of the liposomes. A synthetic 23 amino acid peptide representing the N-terminal sequence of rhodanese was very efficient at disrupting the liposomes. Shorter peptides chosen from within this sequence (residues 11-23 or residues 1-17) had no effect on liposome disruption. A peptide representing the tether region that connects the domains of the enzyme was also without effect. These results are consistent with the hypothesis that the N-terminal sequence of rhodanese is an uncleaved leader sequence, and can interact with membrane components that are involved in the mitochondrial uptake of this protein.
硫氰酸酶（硫代硫酸盐氰化物硫转移酶；E.C. 2.8.1.1）是一种线粒体酶，导入后未进行加工。我们描述了体外实验，该实验表明部分折叠的硫氰酸酶可与脂质双层相互作用。通过测量硫氰酸酶破坏由磷脂酰丝氨酸组成的小单层囊泡以及释放包裹在脂质体中的6 - 羧基荧光素的能力来监测这种相互作用。通过稀释尿素变性的酶得到的部分折叠的硫氰酸酶能有效地诱导脂质体泄漏。天然硫氰酸酶对脂质体完整性没有影响。在引入脂质体之前，如果给予硫氰酸酶重新折叠或聚集的机会，脂质体破坏程度会逐渐降低。一种代表硫氰酸酶N端序列的合成23氨基酸肽在破坏脂质体方面非常有效。从该序列中选取的较短肽段（第11 - 23位残基或第1 - 17位残基）对脂质体破坏没有影响。一种代表连接该酶结构域的系链区域的肽也没有作用。这些结果与以下假设一致：硫氰酸酶的N端序列是一个未切割的前导序列，并且可以与参与该蛋白质线粒体摄取的膜成分相互作用。
