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Human Neuraminidase Isoenzymes Show Variable Activities for 9-O-Acetyl-sialoside Substrates
ACS Chemical Biology ( IF 3.5 ) Pub Date : 2018-01-17 00:00:00 , DOI: 10.1021/acschembio.7b00952 Carmanah D. Hunter 1 , Neha Khanna 1 , Michele R. Richards 1 , Reza Rezaei Darestani 1 , Chunxia Zou 1 , John S. Klassen 1 , Christopher W. Cairo 1
ACS Chemical Biology ( IF 3.5 ) Pub Date : 2018-01-17 00:00:00 , DOI: 10.1021/acschembio.7b00952 Carmanah D. Hunter 1 , Neha Khanna 1 , Michele R. Richards 1 , Reza Rezaei Darestani 1 , Chunxia Zou 1 , John S. Klassen 1 , Christopher W. Cairo 1
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Recognition of terminal sialic acids is central to many cellular processes, and structural modification of sialic acid can disrupt these interactions. A prominent, naturally occurring, modification of sialic acid is 9-O-acetylation (9-O-Ac). Study of this modification through generation and analysis of 9-O-Ac sialosides is challenging because of the lability of the acetate group. Fundamental questions regarding the role of 9-O-Ac sialic acids remain unanswered, including what effect it may have on recognition and hydrolysis by the human neuraminidase enzymes (hNEU). To investigate the substrate activity of 9-O-acetylated sialic acids (Neu5,9Ac2), we synthesized an acetylated fluorogenic hNEU substrate 2′-(4-methylumbelliferyl)-9-O-acetyl-α-d-N-acetylneuraminic acid. Additionally, we generated a panel of octyl sialyllactosides containing modified sialic acids including variation in linkage, 9-O-acetylation, and C-5 group (Neu5Gc). Relative rates of substrate cleavage by hNEU were determined using fluorescence spectroscopy and electrospray ionization mass spectrometry. We report that 9-O-acetylation had a significant, and differential, impact on sialic acid hydrolysis by hNEU with general substrate tolerance following the trend of Neu5Ac > Neu5Gc ≫ Neu5,9Ac2 for NEU2, NEU3, and NEU4. Both NEU2 and NEU3 had remarkably reduced activity for Neu5,9Ac2 containing substrates. Other isoenzymes appeared to be more tolerant, with NEU4 even showing increased activity on Neu5,9Ac2 substrates with an aryl aglycone. The impact of these minor structural changes to sialic acid on hNEU activity was unexpected, and these results provide evidence of the substantial influence of 9-O-Ac modifications on hNEU enzyme substrate specificity. Furthermore, these findings may implicate hNEU in processes governed by 9-O-acetyltransferases and -esterases.
中文翻译：
人类神经氨酸酶同工酶显示9 - O-乙酰唾液酸底物的可变活性
末端唾液酸的识别是许多细胞过程的核心，唾液酸的结构修饰可以破坏这些相互作用。唾液酸的一种明显的，天然存在的修饰是9- O-乙酰化（9- O- Ac）。由于乙酸酯基团的不稳定性，通过生成和分析9- O- Ac唾液酸苷进行这种修饰的研究具有挑战性。关于9- O- Ac唾液酸的作用的基本问题仍未得到解答，包括其对人神经氨酸酶（hNEU）的识别和水解可能产生何种作用。调查9- O-乙酰化唾液酸（Neu5,9Ac 2），我们合成了乙酰化的荧光hNEU底物2'-（4-甲基伞形酮）-9 - O-乙酰基-α - d - N-乙酰基神经氨酸。此外，我们生成了一组包含改性唾液酸的辛基唾液酸乳糖苷，包括键合，9- O-乙酰化和C-5基团（Neu5Gc）的变化。使用荧光光谱法和电喷雾电离质谱法测定hNEU裂解底物的相对速率。我们报道9- O-乙酰化对hNEU的唾液酸水解具有显着的差异影响，具有一般底物耐受性，遵循Neu5Ac> Neu5Gc≫ Neu5,9Ac 2的趋势用于NEU2，NEU3和NEU4。NEU2和NEU3对含Neu5,9Ac 2的底物的活性均显着降低。其他同工酶似乎更具耐受性，NEU4甚至对带有芳基糖苷配基的Neu5,9Ac 2底物显示出更高的活性。这些对唾液酸的微小结构变化对hNEU活性的影响是出乎意料的，并且这些结果提供了9- O- Ac修饰对hNEU酶底物特异性的实质影响的证据。此外，这些发现可能暗示hNEU参与由9- O-乙酰基转移酶和-酯酶控制的过程。
更新日期：2018-01-17
中文翻译：
人类神经氨酸酶同工酶显示9 - O-乙酰唾液酸底物的可变活性
末端唾液酸的识别是许多细胞过程的核心，唾液酸的结构修饰可以破坏这些相互作用。唾液酸的一种明显的，天然存在的修饰是9- O-乙酰化（9- O- Ac）。由于乙酸酯基团的不稳定性，通过生成和分析9- O- Ac唾液酸苷进行这种修饰的研究具有挑战性。关于9- O- Ac唾液酸的作用的基本问题仍未得到解答，包括其对人神经氨酸酶（hNEU）的识别和水解可能产生何种作用。调查9- O-乙酰化唾液酸（Neu5,9Ac 2），我们合成了乙酰化的荧光hNEU底物2'-（4-甲基伞形酮）-9 - O-乙酰基-α - d - N-乙酰基神经氨酸。此外，我们生成了一组包含改性唾液酸的辛基唾液酸乳糖苷，包括键合，9- O-乙酰化和C-5基团（Neu5Gc）的变化。使用荧光光谱法和电喷雾电离质谱法测定hNEU裂解底物的相对速率。我们报道9- O-乙酰化对hNEU的唾液酸水解具有显着的差异影响，具有一般底物耐受性，遵循Neu5Ac> Neu5Gc≫ Neu5,9Ac 2的趋势用于NEU2，NEU3和NEU4。NEU2和NEU3对含Neu5,9Ac 2的底物的活性均显着降低。其他同工酶似乎更具耐受性，NEU4甚至对带有芳基糖苷配基的Neu5,9Ac 2底物显示出更高的活性。这些对唾液酸的微小结构变化对hNEU活性的影响是出乎意料的，并且这些结果提供了9- O- Ac修饰对hNEU酶底物特异性的实质影响的证据。此外，这些发现可能暗示hNEU参与由9- O-乙酰基转移酶和-酯酶控制的过程。
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