Modification and modificatory kinetics of the active center of prawn beta-N-acetyl-D-glucosaminidase.
Beta-N-acetyl-D-glucosaminidase (NAGase, EC3.2.1.52) plays important role in molting, digestion of chitinous foods, and defense systems against parasites in prawn (Litopenaeus vannamei). However, study on functional groups and catalytic mechanism of NAGase are yet limited. The modification of the active center of NAGase from prawn has been first studied. The results demonstrate that the disulfide bonds and the carbamidine groups of arginine residues are not essential to the enzyme's activity. The modification of indole group of tryptophan of the enzyme by N-bromosuccinimide (NBS) can lead to the complete inactivation, accompanying the absorption decreasing at 276 nm, indicating that tryptophan is essential residue to the enzyme. The modificatory kinetics of NAGase in the appropriate concentrations of NBS solution has been studied and the numbers of essential tryptophan residues have been determined using the kinetic method of the substrate reaction. The result shows that only one tryptophan residue is essential for enzyme activity. And the modifications of histidine, lysine residue, and the carboxyl groups also inactivate the enzyme completely or incompletely. The results showed that the carboxyl groups of acidic amino acid, imidazole groups of histidine residue, amino groups of lysine residue, and indole group of tryptophan were essential for the activity of enzyme.
Xie
Xiao-Lan
XL
Department of Oceanography, School of Oceanography and Environmental Sciences, Xiamen University, Xiamen 361005, China.
Huang
Qian-Sheng
QS
Wang
Ye
Y
Ke
Cai-Huan
CH
Chen
Qing-Xi
QX
D000118:Acetylglucosaminidase / Q000737:chemistry / Q000378:metabolism*; D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D001120:Arginine / Q000737:chemistry / Q000378:metabolism; D001974:Bromosuccinimide / Q000737:chemistry / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D006639:Histidine / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D033561:Penaeidae / Q000201:enzymology*; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism
D000596:Amino Acids; D004220:Disulfides; D006639:Histidine; D014364:Tryptophan; D001120:Arginine; D000118:Acetylglucosaminidase; D001974:Bromosuccinimide