Department of Medicinal Chemistry, University of Minnesota, Minneapolis, MN, USA.
Department of Biochemistry Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN, USA.
FEBS Lett. 2020 May;594(10):1497-1505. doi: 10.1002/1873-3468.13745. Epub 2020 Feb 17.
Human histidine triad nucleotide-binding protein 2 (hHINT2) is an important player in human mitochondrial bioenergetics, but little is known about its catalytic capabilities or its nucleotide phosphoramidate prodrug (proTide)-activating activity akin to the cytosolic isozyme hHINT1. Here, a similar substrate specificity profile (k /K ) for model phosphoramidate substrates was found for hHINT2 but with higher k and K values when compared with hHINT1. A broader pH range for maximum catalytic activity was determined for hHINT2 (pK = 6.76 ± 0.16, pK = 8.41 ± 0.07). In addition, the known hHINT1-microphthalmia-inducing transcription factor-regulating molecule Ap A was found to have no detectable binding to HINT1 nor HINT2 by isothermal titration calorimetry. These results demonstrate that despite differences in their sequence and localization, HINT1 and HINT2 have similar nucleotide substrate specificities, which should be considered in future proTide design and in studies of their natural function.
人组氨酸三联核苷酸结合蛋白 2（hHINT2）是人类线粒体生物能量学的重要参与者，但对其催化能力或与其类似的胞质同工酶 hHINT1 类似的核苷酸磷酰胺前药（proTide）激活活性知之甚少。在这里，发现 hHINT2 对模型磷酰胺底物具有相似的底物特异性谱（k / K ），但与 hHINT1 相比，k 和 K 值更高。确定 hHINT2 的最大催化活性具有更宽的 pH 范围（pK = 6.76 ± 0.16，pK = 8.41 ± 0.07）。此外，通过等温滴定量热法发现，已知的 hHINT1-小眼畸形诱导转录因子调节分子 Ap A 对 HINT1 和 HINT2 均无检测到的结合。这些结果表明，尽管 HINT1 和 HINT2 在序列和定位上存在差异，但它们具有相似的核苷酸底物特异性，这在未来的 proTide 设计和对其天然功能的研究中应加以考虑。
