Enzymatic and molecular properties of the Clostridium tertium sialidase.
Clostridium tertium metabolizes sialoglycoconjugates via a secreted sialidase [EC 3.2.1.18] and an intracellular acylneuraminate pyruvate lyase [EC 4.1.3.3]. The sialidase was enriched 1,900-fold from the culture medium with a specific activity of 0.7 U per mg protein. It exhibits a temperature optimum of 50 degreesC and tolerates mercury ions at relatively high concentrations (50% inhibition at 5.2 mM Hg2+). The sialidase gene was detected on two restriction fragments (HincII, HindIII) of chromosomal DNA and their correct recombination resulted in an active enzyme expressed by Escherichia coli. The structural gene is represented by 2,319 bp encoding a protein of 773 amino acids with a molecular mass of 85.4 kDa. The first 28 amino acids possess the character of a signal peptide. The protein reveals a FRIP-region and four Asp-boxes common in all bacterial sialidases. It has 42.6% identical amino acids when compared with the sialidase of Clostridium septicum and 64.8% with a sialidase gene amplified from Macrobdella decora. A further open reading frame was detected 30 bp downstream from the sialidase gene, which exhibits significant homology with acylneuraminate pyruvate lyases. For the first time, both genes were found in close proximity on a bacterial chromosome, probably being part of one operon.
Grobe
K
K
Biochemisches Institut, Christian-Albrechts-Universität, D-24098, Kiel, Germany.
Sartori
B
B
Traving
C
C
Schauer
R
R
Roggentin
P
P
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology*; D008628:Mercury / Q000494:pharmacology; D008969:Molecular Sequence Data; D009439:Neuraminidase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D007652:Oxo-Acid-Lyases / Q000235:genetics / Q000378:metabolism; D010802:Phylogeny; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid
D011994:Recombinant Proteins; D009439:Neuraminidase; D007652:Oxo-Acid-Lyases; C018728:N-acetylneuraminate lyase; D008628:Mercury