Department of Pharmacology and Physiology, UMDNJ, New Jersey Medical School, Newark, NJ 07101, United States.
Mitochondrion. 2012 Sep;12(5):539-49. doi: 10.1016/j.mito.2012.07.103. Epub 2012 Jul 17.
Cysteine desulfurases generate a covalent persulfide intermediate from cysteine, and this activated form of sulfur is essential for the synthesis of iron-sulfur (Fe-S) clusters. In yeast mitochondria, there is a complete machinery for Fe-S cluster synthesis, including a cysteine desulfurase, Nfs1p. Here we show that following supplementation of isolated mitochondria with [(35)S]cysteine, a radiolabeled persulfide could be detected on Nfs1p. The persulfide persisted under conditions that did not permit Fe-S cluster formation, such as nucleotide and/or iron depletion of mitochondria. By contrast, under permissive conditions, the radiolabeled Nfs1p persulfide was greatly reduced and radiolabeled aconitase was formed, indicating transfer of persulfide to downstream Fe-S cluster recipients. Nfs1p in mitochondria was found to be relatively more resistant to inactivation by N-ethylmaleimide (NEM) as compared with a prokaryotic cysteine desulfurase. Mitochondria treated with NEM (1 mM) formed the persulfide on Nfs1p but failed to generate Fe-S clusters on aconitase, likely due to inactivation of downstream recipient(s) of the Nfs1p persulfide. Thus the Nfs1p-bound persulfide as described here represents a precursor en route to Fe-S cluster synthesis in mitochondria.
半胱氨酸脱硫酶从半胱氨酸中生成一个共价过硫化物中间体，这种活化形式的硫对于铁硫（Fe-S）簇的合成至关重要。在酵母线粒体中，存在一个完整的 Fe-S 簇合成机制，包括一种半胱氨酸脱硫酶，Nfs1p。在这里，我们表明，在用 [(35)S]半胱氨酸补充分离的线粒体后，可以在 Nfs1p 上检测到放射性标记的过硫化物。在不允许形成 Fe-S 簇的条件下，例如线粒体核苷酸和/或铁耗竭，过硫化物仍然存在。相比之下，在允许的条件下，放射性标记的 Nfs1p 过硫化物大大减少，形成了放射性标记的 aconitase，表明过硫化物转移到下游 Fe-S 簇受体。与原核半胱氨酸脱硫酶相比，在线粒体中，Nfs1p 相对更能抵抗 N-乙基马来酰亚胺（NEM）的失活。用 NEM（1 mM）处理的线粒体在 Nfs1p 上形成过硫化物，但未能在 aconitase 上生成 Fe-S 簇，可能是由于 Nfs1p 过硫化物下游受体的失活。因此，这里描述的 Nfs1p 结合的过硫化物代表了在线粒体中 Fe-S 簇合成过程中的前体。
