Inhibitory kinetics of betaine on beta-N-acetyl-D-glucosaminidase from prawn (Litopenaeus vannamei).
The effects of betaine on prawn beta-N-acetyl-D-glucosaminidase (NAGase) activity for the hydrolysis of p-nitrophenyl-N-acetyl- beta-D-glucosaminide (pNP-NAG) have been studied. The results showed that appropriate concentrations of betaine could lead to reversible inhibition against NAGase, and the IC(50) value was estimated to be 15.00 +/- 0.30 mM. The inhibitory kinetics assay showed that betaine was a mixed type inhibitor with a K(I) value of 9.17 +/- 0.85 mM and a K(IS) value of 45.58 +/- 2.52 mM. The inhibitory model was set, and the microscopic rate constants were determined using the kinetic method of the substrate reaction. The time course of the hydrolysis of pNP-NAG catalyzed by NAGase in the presence of different betaine concentrations showed that at each betaine concentration, the rate decreased with an increase in time until a straight line was approached, indicating that the inhibition of NAGase by betaine is a slow, reversible reaction with fractional residual activity. The fact that k(+0) is much larger than k(+0)(') indicated that the free enzyme molecule is more fragile than the enzyme-substrate complex against betaine. It is suggested that the presence of the substrate offers marked protection of NAGase against inhibition by betaine.
Xie
Xiao-Lan
XL
Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystems, School of Life Sciences, Xiamen University, Xiamen, China.
Zhou
Han-Tao
HT
Huang
Qian-Sheng
QS
Wei
Xiao-Qian
XQ
Wang
Ye
Y
Yan
Jiang-Hua
JH
Chen
Qing-Xi
QX
D000118:Acetylglucosaminidase / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D000818:Animals; D001622:Betaine / Q000737:chemistry*; D003445:Crustacea / Q000737:chemistry / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000737:chemistry*; D007700:Kinetics; D011485:Protein Binding
D004791:Enzyme Inhibitors; D001622:Betaine; D000118:Acetylglucosaminidase