Abstract
Three site-specific mutations were performed in two regions of a sialidase gene fromClostridium perfringens which are known to be conserved in bacterial sialidases. The mutant enzymes were expressed inEscherichia coli and, when measured with MU-Neu5Ac as substrate, exhibited variations in enzymatic properties compared with the wild-type enzyme. The conservative substitution of Arg 37 by Lys, located in a short conserved region upstream from the four repeated sequences common in bacterial sialidase genes, was of special interest, asK M andV max, as well asK i measured with Neu5Ac2en, were dramatically changed. These data suggest that this residue may be involved in substrate binding. In addition to its low activity, this mutant enzyme has a lower temperature optimum and is active over a more limited pH range. This mutation also prevents the binding of an antibody able to inhibit the wild-type sialidase. The other mutations, located in one of the consensus sequences, were of lower influence on enzyme activity and recognition by antibodies.
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Roggentin, T., Kleineidam, R.G., Schauer, R. et al. Effects of site-specific mutations on the enzymatic properties of a sialidase fromClostridium perfringens . Glycoconjugate J 9, 235–240 (1992). https://doi.org/10.1007/BF00731135
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DOI: https://doi.org/10.1007/BF00731135