扫描丙氨酸白色念珠菌的诱变和de-peptidization myristoyl-CoA:蛋白质N-myristoyltransferase八肽底物显示三个元素分子识别的关键。
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McWherter CA、Rocque WJ Zupec我,弗里曼SK,布朗DL Devadas B -盖特曼DP,西科尔斯基是的,戈登霁
扫描丙氨酸白色念珠菌的诱变和de-peptidization myristoyl-CoA:蛋白质N-myristoyltransferase八肽底物显示三个元素分子识别的关键。
生物化学杂志。1997年5月2日,272 (18):11874 - 80。
- PubMed ID
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9115247 (在PubMed]
- 文摘
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白色念珠菌产生一个myristoyl-CoA:蛋白质N-myristoyltransferase (Nmt)对其生存能力至关重要。(Arf)是一个ADP-ribosylation因素包括少数细胞蛋白质底物的酶。一个八肽(GLYASKLS-NH2)来自一个氨基端Arf序列作为出发点来标识元素识别酰基转移酶的peptide-binding网站的关键。体外动力学研究中,采用纯化Nmt和一组多肽与单一Ala替换GLYASKLS-NH2每个位置,证实其Gly1, Ser5, Lys6残留物中扮演主要的角色绑定。ALYASKLS-NH2被发现的抑制剂竞争肽(Ki = 15.3 + / - 6.4 microM)和非竞争性myristoyl-CoA (Ki = 31.2 + / - 0.7 microM)。调查26衍生品的抑制剂,(我)一个完整的丙氨酸扫描,(ii)进步c端截断,和(3)操纵的理化特性和它的残留1 5和6,证实了重要的氨基胺立体化学的要求,Ser5 beta-hydroxyl,和Lys6 epsilon-amino组。值得注意的是,取代氨基四肽的ALYASKLS-NH2 11-aminoundecanoyl集团生产的竞争性抑制剂,11-aminoundecanoyl-SKLS-NH2, 38倍更有效(Ki = 0.40 + / - 0.03 microM)比开始八肽。消除了伯胺(undecanoyl-SKLS-NH2),或与一个甲基取代它(dodecanoyl-SKLS-NH2),导致26 - 34倍增加IC50,确认胺的重要贡献的认可。从C末端切除LeuSer (11-aminoundecanoyl-SK-NH2)产生了竞争二肽抑制剂与Ki (11.7 + / - 0.4 microM)相当于开始八肽,ALYASKLS-NH2。替换的Ser高丝氨酸、cis-4-hydroxyproline或酪氨酸减少效力的3 - 70倍,强调要求适当的演讲中羟基的二肽抑制剂。 Substituting D- for L-Lys decreases its inhibitory activity >100-fold, while deletion of the epsilon-amino group (Nle) or masking its charge (epsilon-N-acetyl-lysine) produces 4-7-fold attenuations. L-His, but not its D-isomer, can fully substitute for L-Lys, producing a competitive dipeptide inhibitor with similar potency (Ki = 11.9 +/- 1.0 microM). 11-Aminoundecanoyl-SK-NH2 and 11-aminoundecanoyl-SH-NH2 establish that a simple alkyl backbone can maintain an appropriate distance between three elements critical for recognition by the fungal enzyme's peptide-binding site: a simple omega-terminal amino group, a beta-hydroxyl, and an epsilon-amino group or an imidazole. These compounds contain one peptide bond and two chiral centers, suggesting that it may be feasible to incorporate these elements of recognition, or functionally equivalent mimics, into a fully de-peptidized Nmt inhibitor.