Matovina Mihaela, Tomašić Paić Ana, Tomić Sanja, Brkić Hrvoje, Horvat Lucija, Barbarić Lea, Filić Vedrana, Pinterić Marija, Jurić Snježana, Kussayeva Akmaral
Division of Organic Chemistry and Biochemistry, Ruđer Bošković Institute, Bijenička Cesta 54, 10000 Zagreb, Croatia.
Faculty of Medicine, Josip Juraj Strossmayer University of Osijek, 31000 Osijek, Croatia.
Int J Mol Sci. 2023 Sep 16;24(18):14178. doi: 10.3390/ijms241814178.
Dipeptidyl peptidase 3 (DPP3) is a zinc-dependent exopeptidase with broad specificity for four to eight amino acid residue substrates. It has a role in the regulation of oxidative stress response NRF2-KEAP1 pathway through the interaction with KEAP1. We have conducted stable isotope labeling by amino acids in a cell culture coupled to mass spectrometry (SILAC-MS) interactome analysis of TRex HEK293T cells using DPP3 as bait and identified SH2 Domain-Containing Protein 3C (SH2D3C) as prey. SH2D3C is one of three members of a family of proteins that contain both the SH2 domain and a domain similar to guanine nucleotide exchange factor domains of Ras family GTPases (Ras GEF-like domain), named novel SH2-containing proteins (NSP). NSPs, including SH2D3C (NSP3), are adaptor proteins involved in the regulation of adhesion, migration, tissue organization, and immune response. We have shown that SH2D3C binds to DPP3 through its C-terminal Ras GEF-like domain, detected the colocalization of the proteins in living cells, and confirmed direct interaction in the cytosol and membrane ruffles. Computational analysis also confirmed the binding of the C-terminal domain of SH2D3C to DPP3, but the exact model could not be discerned. This is the first indication that DPP3 and SH2D3C are interacting partners, and further studies to elucidate the physiological significance of this interaction are on the way.
二肽基肽酶3（DPP3）是一种锌依赖性外肽酶，对四至八个氨基酸残基底物具有广泛的特异性。它通过与KEAP1相互作用，在氧化应激反应NRF2-KEAP1途径的调节中发挥作用。我们利用DPP3作为诱饵，对TRex HEK293T细胞进行了细胞培养中氨基酸稳定同位素标记结合质谱（SILAC-MS）相互作用组分析，并鉴定出含SH2结构域蛋白3C（SH2D3C）为猎物。SH2D3C是一类蛋白质的三个成员之一，这类蛋白质既含有SH2结构域，又含有一个与Ras家族GTP酶的鸟嘌呤核苷酸交换因子结构域相似的结构域（Ras GEF样结构域），被称为新型含SH2蛋白（NSP）。包括SH2D3C（NSP3）在内的NSP是参与调节黏附、迁移、组织构建和免疫反应的衔接蛋白。我们已经表明，SH2D3C通过其C末端的Ras GEF样结构域与DPP3结合，检测到了这两种蛋白在活细胞中的共定位，并证实了它们在细胞质和膜褶皱中的直接相互作用。计算分析也证实了SH2D3C的C末端结构域与DPP3的结合，但具体模型尚不清楚。这是DPP3和SH2D3C为相互作用伙伴的首个迹象，进一步阐明这种相互作用生理意义的研究正在进行中。
