Isolation and properties of the natural and the recombinant sialidase fromClostridium septicum NC 0054714 |
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Authors: | Kirsten I Zenz Peter Roggentin Roland Schauer |
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Institution: | (1) Biochemisches Institut, Christian-Albrechts-Universität, Olshausenstr, 40, D-2300 Kiel, Germany |
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Abstract: | The natural sialidase ofClostridium septicum was purified and characterized in parallel with the recombinant enzyme expressed byEscherichia coli. The two enzymes exhibit almost identical properties. The maximum hydrolytic activity was measured at 37 °C in 60mm sodium acetate buffer, pH 5.3. Glycoproteins like fetuin and saponified bovine submandibular gland mucin, most of them having (2-6) linked sialic acids, are preferred substrates, while sialic acids from gangliosides, sialyllactoses, or the (2-8) linked sialic acid polymer (colominic acid) are hydrolysed at lower rates. (2-3) Linkages are more rapidly hydrolysed than (2-6) bonds of sialyllactoses. The cleavage rate is markedly reduced by O-acetylation of the sialic acid moiety. These properties are similar to those of other secreted clostridial sialidases. The enzyme exists in mono-, di- and trimeric forms, the monomer exhibiting a molecular mass of 125 kDa, which is close to the protein mass of 111 kDa deduced from the nucleotide sequence of the cloned gene.Abbreviations BSM
bovine submandibular gland mucine
- CMM
cooked meat medium
- EDTA
ethylenediaminetetraacetic acid
- FPLC
fast performance liquid chromatography
- LB
Luria-Bertani
- MU-Neu5Ac
4-methylumbelliferyl- -d-N-acetylneuraminic acid
- Neu5Ac
N-acetylneuraminic acid
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid
- Neu4,5Ac2
N-acetyl-4-O-acetylneuraminic acid
- pI
isoelectric point
- SDS
sodium dodecyl sulfate |
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Keywords: | sialidase (neuraminidase) cloned and natural enzyme isolation comparison of properties Clostridium septicum |
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