Small Sialidases

Chang, Y. K., and Chein, C. H. (1998). Simple two step procedure for purification of cloned small sialidase from unclaridied E. coli feedstocks, lnt. Conf. Expanded Bed Adsorption, 2nd, Napa Valley, CA, 1998, Abstr., p. 8.2. 

Starting with the crystal structure of the bacterial sialidase of Salmonella typhi-murium [60] which is used as a framework, the knowledge based homology modelling produces a set of energy-minimised conformations for the small sialidase of C. 

Figure 2. The sialidase superfamily (a) Influenza virus neuraminidase tetramer, (b) the small sialidases from S. typhimurium and (c) M. viridifaciens, (d) the side view of the large M. viridifaciens sialidase, (e) V. cholerae sialidase. The inhibitor Neu5Ac2en is shown bound in several active sites and galactose is also shown in the large M. viridifaciens sialidase. The isolated grey spheres represent divalent ions that play a key structural role in influenza and V. cholerae sialidase, and are an absolute requirement for activity.

The crystal structures of two representative small sialidases, with molecular weights around 40 kDa, have been determined one from Salmonella typhimurium [17] and one from Micromonospora viridifaciens [I8j. These reveal the same P-propeller fold seen in the influenza virus neuraminidase (Figure 2), despite having no sequence similarity to the viral enzyme, and not containing any disulphide bonds in contrast to the seven conserved disulfides in the viral enzyme. 

Prepare a fresh fibroblast homogenate in 130 pi distilled water (note sialidase is highly labile upon sonification and freezing ), using a small (200-pl) Potter appara-... 

Marine sponges contain a host of bioactive compounds, particularly small molecules, and also contain a range of peptides that are non-ribosomally synthesised, often containing non-native amino acids. However, there are examples of peptides of ribosomal origin, including, for example, asteropine A isolated from the sponge Asteropus simplex.133 This peptide comprises 36 residues and three disulphide bonds. It has potent sialidase inhibitory activity and thus has applications in the design of novel viral inhibitors. Structural analysis of asteropine A with NMR spectroscopy revealed a cystine-knot motif, similar to that already described for plant toxins. This observation emphasises the fact that the cystine-knot motif is extremely prevalent in disulphide-rich peptides.134 Asteropine A, discovered in 2006, was the first reported cystine-knot peptide isolated from marine invertebrates other than from cone snails, which are described in more detail below. 

T. cruzi small mucin-like gene family TcTS, Trypanosoma cruzi trans-sialidase THF, tetrahydrofuran TMSOTf, trimethylsilyl trifluoromethanesulfonate TRR, Thr-rich region TSSA, trypomastigote small surface antigen UDP, uridine diphosphate VSG, variant surface glycoprotein. 

Although there is an occasional recruitment of metal ions as structurally stabilising elements e.g. GH 35 sialidases require Ca " ) or metals and small organic molecules as part of the acid/base machinery, except for GH 4 enzymes there are no cofactor requirements catalysis is wholly carried out by the protein itself. 

This Neu5Ac derivative lacking a glycosidic hydroxyl group at C2 has been found as free sialic acid in small quantities in blood serum, saliva and urine of man [5,8,13] and in tissue extracts of starfish [36] (Table 1). It is unknown, whether this compound, which is a potent competitive inhibitor of sialidases (see below), has a biological function. Its formation under physiological conditions has been described in section 2. 

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