N-acetyl alpha-D-glucosaminidase from the venom of African puff adder (Bitis arietans).
The activity of N-acetyl-alpha-D-glucosaminidase from venom of the African puff adder (Bitis arietans) has been detected. The enzyme from the venom was purified by chromatography on Q-sepharose, CM-cellulose, and N-acetyl-alpha-D-glucosamine-agarose affinity column. The enzyme has a molecular weight of 102 kDa determined by size exclusion chromatography on Sephacryl 200. It migrated as a 51-kDa band on SDS polyacrylamide gels. The enzyme is maximally active at pH 5.5 and 40 degrees C. The B. arietans NAGase hydrolyzed exclusively terminally linked alpha-(1-4) GlcNAc residues from nonreducing ends of oligosaccharides. It hydrolysed chito-oligosaccharide, MU-GlcNAc and chitobiose with K(M) values of 0.15 mM and 1.22 mM, respectively. Swollen chitin and oligosaccharide above (GlcNAc)(4) were not hydrolysed by the enzyme. B. arietans NAGase was strongly inhibited noncompetitively by Hg(2+), competitively by 1-thio-beta-D-GlcNAc and N-acetyl glucosamine (NAG) with K(i) of 0.55, 0.25 and 8 mM, respectively. Colombin the active component of antivenom preparation from Aristolodia albida inhibited the enzyme competitively with K(i) of 0.6 mM. Delineation of the active site by chemical modification revealed the involvement of His and Trp in the catalysis of the enzyme.
Nok
A J
AJ
Department of Biochemistry, Ahmadu Bello University, Zaria, Nigeria.
Shuaibu
M N
MN
Choudhry
M K
MK
Oyebanjo
O
O
Ibrahim
S
S
Williams
S
S
D000118:Acetylglucosaminidase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000633:toxicity; D001665:Binding Sites; D002846:Chromatography, Affinity; D004355:Drug Stability; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006639:Histidine / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D013379:Substrate Specificity; D013696:Temperature; D014757:Viper Venoms / Q000201:enzymology*
D004791:Enzyme Inhibitors; D014757:Viper Venoms; D006639:Histidine; C033079:alpha-N-acetyl-D-glucosaminidase; D000118:Acetylglucosaminidase