可光活化的 O-GlcNAc 转移酶文库可对溶剂暴露的 TPR 结构域相互作用进行共价化学捕获。

IF 2.6 4区 生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY
ChemBioChem Pub Date : 2024-11-14 DOI:10.1002/cbic.202400709
Cassandra M Joiner, Tiarra J Glogowski, Erin M NewRingeisen, Huy V Huynh, Melanie G Roberts, Madison M Rognerud, Hahns E Huebsch
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引用次数: 0

摘要

O- 链接 N-乙酰葡糖胺(O-GlcNAc)转移酶(OGT)是一种重要的应激感应酶,负责将 O-GlcNAc 单糖添加到数千种核蛋白质和细胞质蛋白质中,以调节细胞的平衡。OGT 底物几乎存在于所有细胞内过程中,蛋白质 O-GlcNAc 水平的扰动与蛋白质静态疾病(如癌症、代谢紊乱和神经变性)有关。这种广泛的疾病活性使 OGT 成为一个有吸引力的治疗靶点;然而,底物的多样性使泛抑制作为一种治疗策略变得不可行。相反,底物特异性靶向方法更具优势,但人们对 OGT 如何选择其底物仍知之甚少。底物特异性是由 OGT 的非催化四肽重复(TPR)结构域而非其糖基转移酶结构域之间的相互作用控制的。OGT 的 TPR 结构域形成了一个 100 Å 的超螺旋结构,包含一个称为底物结合表面的内腔表面和一个溶剂暴露表面。迄今为止,还没有工具能对该结构域的区域进行定点选择,并区分两个结合表面。在这里,我们开发了一个重组 OGT 构建库,其中含有沿 TPR 结构域溶剂暴露表面定点结合的可光激活的非天然氨基酸 (UAAs),以共价捕获和绘制 OGT 的相互作用组。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Photoactivatable O-GlcNAc Transferase Library Enables Covalent Chemical Capture of Solvent-Exposed TPR Domain Interactions.

O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) is an essential, stress-sensing enzyme responsible for adding the O-GlcNAc monosaccharide to thousands of nuclear and cytoplasmic proteins to regulate cellular homeostasis. OGT substrates are found in almost all intracellular processes, and perturbations in protein O-GlcNAc levels have been implicated in proteostatic diseases, such as cancers, metabolic disorders, and neurodegeneration. This broad disease activity makes OGT an attractive therapeutic target; however, the substrate diversity makes pan-inhibition as a therapeutic strategy unfeasible. Rather, a substrate-specific approach to targeting is more advantageous, but how OGT chooses its substrates remains poorly understood. Substrate specificity is controlled by the interactions between OGT's non-catalytic tetratricopeptide repeat (TPR) domain, rather than its glycosyltransferase domain. OGT's TPR domain forms a 100 Å superhelical structure, containing a lumenal surface, known as the substrate-binding surface, and a solvent-exposed surface. To date, there are no tools to site-selectively target regions of the domain and differentiate between the two binding surfaces. Here, we developed a library of recombinant OGT constructs containing site-specifically incorporated photoactivatable unnatural amino acids (UAAs) along the solvent-exposed surface of the TPR domain to covalently capture and map OGT's interactome.

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来源期刊
ChemBioChem
ChemBioChem 生物-生化与分子生物学
CiteScore
6.10
自引率
3.10%
发文量
407
审稿时长
1 months
期刊介绍: ChemBioChem (Impact Factor 2018: 2.641) publishes important breakthroughs across all areas at the interface of chemistry and biology, including the fields of chemical biology, bioorganic chemistry, bioinorganic chemistry, synthetic biology, biocatalysis, bionanotechnology, and biomaterials. It is published on behalf of Chemistry Europe, an association of 16 European chemical societies, and supported by the Asian Chemical Editorial Society (ACES).
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