Keywords: 
• Hg²⁺ /
• Pb²⁺ /
• Tachypleus tridentatus /
• NAGase /
• inhibition /
• conformational change

Abstract: The molting of arthropods is closely related to N-acetyl-β-D-glucosaminidase (EC3.2.1. 52, NAGase). In order to investigate the effects of Hg²⁺ and Pb²⁺ heavy metal ions on NAGase from arthropods, the NAGase were extracted from the viscera of Tachypleus tridentatus. The effects of Hg²⁺ and Pb²⁺ on NAGase were studied by using the kinetics method of enzymatic reaction. Besides, the conformational changes of NAGase caused by Hg²⁺ and Pb²⁺ treatment were evaluated by the enzyme fluorescence emission spectrometry, respectively. The results showed that both Hg²⁺ and Pb²⁺ had strong inhibitory effects on the NAGase. The inhibitory effect of Hg²⁺ on NAGase was stronger than that of Pb²⁺. The inhibitory effects of Hg²⁺ and Pb²⁺ on NAGase were both reversible. The inhibitory effect of Hg²⁺ on NAGase was the competitive type and its inhibition constant K_I was 22.68 μmol·L^-1. Hg²⁺ only combined with the free enzymes (E) and did not combine with the enzyme-substrate complex (ES). However, the inhibitory effect of Pb²⁺ on NAGase was the competitive and non-competitive mixed type. The inhibition constants K_I and K_IS were 19.13 mmol·L^-1 and 75.23 mmol·L^-1, respectively. The affinity of Pb²⁺ for free enzyme was stronger than the enzyme-substrate complex. The fluorescence emission intensity of the NAGase treated by Hg²⁺ and Pb²⁺ were both decreased, but their fluorescence emission peak of NAGase did not shift. The results indicated that the inhibitory effects of both Hg²⁺ and Pb²⁺ on NAGase were caused by the conformational changes, which suggested that Hg²⁺ and Pb²⁺ had regulatory effects on the activities of NAGase from Tachypleus tridentatus.