Diversity in the properties of two sialidase isoenzymes produced by Clostridium perfringens spp.
Clostridium perfringens produces two sialidases, one of which has a molecular mass of 71 kDa and is secreted, while the 'small', 43 kDa isoenzyme remains in the cells. The secreted, higher molecular mass sialidases of two different clostridial strains, DSM756T and A99, exhibit maximum activity at pH 5.5 and at 51 or 55 degrees C, respectively. The molecular mass of both enzymes is 71 kDa in SDS-PAGE and 63 kDa as determined by gel-filtration, which indicates the absence of subunits. Natural sialidase substrates are hydrolyzed at comparably high rates, e.g. the glycoproteins fetuin and bovine submandibular gland mucin, the homopolymer colominic acid, and the ganglioside mixture from bovine brain. The partially purified 'small' isoenzyme from C. perfringens A99 cells had similar properties to the corresponding recombinant sialidase isolated from the Escherichia coli host. It is located inside the clostridial and E. coli cells and exhibits maximum activity at pH 6.1 and 37 degrees C. A relative molecular mass of 32,000 was found with FPLC gel-filtration chromatography, while primary structure analysis yielded a value of 43,000. It differs a significantly from the 'large' isoenzyme by substrate specificity. Preferred substrates are oligosaccharides, while other, more complex sialoglycoconjugates are hydrolyzed only at very low rates. alpha 2,3-linkages are hydrolyzed much faster than alpha 2,6-bonds.
Roggentin
P
P
Biochemisches Institut, Christian-Albrechts-Universität, Kiel, Germany.
Kleineidam
R G
RG
Schauer
R
R
D002850:Chromatography, Gel; D003016:Clostridium perfringens / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D009439:Neuraminidase / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis; D013379:Substrate Specificity; D013696:Temperature
D007527:Isoenzymes; D011994:Recombinant Proteins; D009439:Neuraminidase