Enzymatic and Molecular Properties of the Clostridium tertium Sialidase

Abstract

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.

Characterization of Sialidase from an Influenza A (H3N2) Virus Strain: Kinetic Parameters and Substrate Specificity

“…To obtain NA crystals, the utilization of chromatographic methodologies based on charge properties of the sialidase molecule [5,42] has provided preparations ex-pressing one single band; however, they exhibit loss of that enzyme activity which is considered as essential for our purposes [13,36,47].…”

“…The sialidase digestion with pronase introduces a proteolytic contaminant that is not separated by a subsequent purification step as sucrose gradient centrifugation [26,46]. In the ion exchange chromatographic method, a significant loss of sialidase activity is observed [13]. The present paper characterizes an active sialidase purified by pronase treatment and gel filtration, virus structure that was originated from a variant strain of the influenza A virus sample that caused the 1975 pandemic, analyzing some of its properties.…”

Genetic and functional characterization of the NanA sialidase from Clostridium chauvoei

“…Our genetic and biochemical data reveal a sialidase, named NanA, in C. chauvoei that is a 150 kDa homodimer composed of two identical polypeptides of an apparent molecular mass of 72 kDa. Differences between the predicted size calculated from the nucleotide sequence of nanA (81 kDa) and the apparent size observed by SDS-PAGE (72 kDa) have previously been reported for several bacterial sialidases, such as S. pneumoniae [43,44], Clostridium tertium [45] and C. septicum [46]. Native sialidase enzymes from bacteria are described as multimers, mostly dimers, of polypeptides having molecular masses ranging between 50 and 80 kDa [45], which agrees with our data for the sialidase of C. chauvoei .…”

The Leukemoid Reaction inClostridium sordelliiInfection: Neuraminidase Induction of Promyelocytic Cell Proliferation

“…Other clostridial neuraminidases include those produced by C. perfringens (which makes 2) [26], C. septicum [27], C. chauvoei [28] and C. tertium [29]. The C. sordellii neuraminidase is a highly active enzyme that is detectable in wound exudates and serum only hours after infection [30].…”