确定赖氨酸116为铁氧化还原蛋白：NADP⁺氧化还原酶的N-乙基马来酰亚胺失活靶点。

Identification of Lys116 as the target of N-ethylmaleimide inactivation of ferredoxin:NADP+ oxidoreductase.

作者信息

Aliverti A, Gadda G, Ronchi S, Zanetti G

机构信息

Dipartimento di Fisiologia e Biochimica Generali, Università di Milano, Italy.

出版信息

Eur J Biochem. 1991 May 23;198(1):21-4. doi: 10.1111/j.1432-1033.1991.tb15981.x.

DOI:10.1111/j.1432-1033.1991.tb15981.x

PMID:1904024

Abstract

Oxidized ferredoxin:NADP+ oxidoreductase (FNR) was slowly and irreversibly inactivated by N-ethylmaleimide. Complete protection against inactivation was afforded by saturating concentrations of NADP+. In the presence of NADPH, a rapid inhibition of the enzyme ensued; however, this inhibition was found to be reversible. In the tryptic map of the flavoprotein, modified with N-ethyl[2,3-14C]maleimide in oxidizing conditions, a unique radioactive peptide was found. Its sequence comprised residues 110-117 of the enzyme: Lys116 was shown to be the residue alkylated by N-ethylmaleimide. It is noteworthy that the same residue of FNR was found to be modified by 5-dimethylaminoaphthalene-1-sulfonyl(dansyl) chloride at the putative NADP(H)-binding site [Cidaria, D., Biondi, P. A., Zanetti, G. & Ronchi, S. (1985) Eur. J. Biochem. 146, 295-299]. Furthermore, the data reported here demonstrate that the sulfhydryl groups of FNR are not involved in enzyme inactivation by N-ethylmaleimide.

摘要

氧化型铁氧化还原蛋白

NADP⁺氧化还原酶（FNR）被N - 乙基马来酰亚胺缓慢且不可逆地失活。饱和浓度的NADP⁺能完全保护该酶不被失活。在NADPH存在的情况下，会迅速抑制该酶的活性；然而，这种抑制是可逆的。在用N - 乙基[2,3 -¹⁴C]马来酰亚胺在氧化条件下修饰的黄素蛋白的胰蛋白酶图谱中，发现了一个独特的放射性肽段。其序列包含该酶的110 - 117位残基：Lys116被证明是被N - 乙基马来酰亚胺烷基化的残基。值得注意的是，在假定的NADP(H)结合位点，FNR的同一个残基也被5 - 二甲基氨基萘 - 1 - 磺酰氯（丹磺酰氯）修饰过[西达里亚，D.，比昂迪，P. A.，扎内蒂，G. & 龙奇，S.（1985年）《欧洲生物化学杂志》146卷，295 - 299页]。此外，此处报道的数据表明FNR的巯基不参与N - 乙基马来酰亚胺导致的酶失活过程。