Characterization of a sialidase (neuraminidase) isolated from Clostridium chauvoei (Jakari strain).
A sialidase from Clostridium chauvoei (Jakari strain), an indigenous bacterial strain that causes blackleg in Nigerian cattle and other ruminants was isolated and partially purified by chromatography on DEAE cellulose, hydroxyapatite and phenyl agarose columns. The enzyme migrated as a 65-kDa protein after electrophoresis on sodium dodecyl sulphate polyacrylamide gels. It was optimally active at pH 4.5 and 40 degrees C with an activation energy (Ea) of 13.40 kJ mol(-1). It had Km and Vmax values of 170 microM and 200 micromole h(-1) mg(-1) respectively with fetuin as substrate. When sialyllactose (Neu5Ac2,3 lactose) was used as substrate the Km and Vmax values were 8 microM and 5 micromoles min(-1) mg(-1) respectively. The Clostridium chauvoei sialidase cleaved sialic acids from RBC ghosts of sheep, horse, goat, cattle, pig and mice as well as mouse brain cells, albeit at different rates. The enzyme was activated by Ca2+ and Mg2+ and inhibited by the group-specific reagents diethylpyrocarbonate (DEP) and N-ethylmalemide (NEM). The sialidase inhibitors, 2,3 didehydroneuraminic acid (Neu5Ac2,3en) and paranitrophenyl oxamic acid (pNPO) inhibited the enzyme competitively with Ki values of 40 and 30 microM respectively.
Useh
N M
NM
Department of Veterinary Pathology and Microbiology, Ahmadu Bello University, Zaria, Nigeria.
Ajanusi
J O
JO
Esievo
K A N
KA
Nok
A J
AJ
D000818:Animals; D002417:Cattle; D046975:Clostridium chauvoei / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D009438:Neuraminic Acids / Q000378:metabolism; D009439:Neuraminidase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D012794:Sialic Acids / Q000378:metabolism; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D009438:Neuraminic Acids; D012794:Sialic Acids; D009439:Neuraminidase; D019158:N-Acetylneuraminic Acid