Lyase Clostridium perfringens

TV-Acetyl neuraminic acid aldolase [from Clostridium perfringens, TV-acetyIneuraminic acid pyruvate lyase] [9027-60-5] 32,000 [EC 4.1.3.3]. Purified by extraction with H2O,... 

Scheme 4.—Induction and Cooperation of the Sialic acid-specific Enzymes Siali-dase, Acylneuraminate Pyruvate-lyase, and Neu5Ac-permease (hypothetical) in Clostridium perfringens. [Key , sialoglycoprotein , free sialic acid f—at>, ...

A-Acetyl neuraminic acid aldolase [from Clostridium perfringens, A-acetylneuraminic acid pyruvate lyase] [9027-60-5] [EC 4.1.3.3]. Purified by extraction with H20, protamine pptn, (NH4)2S04 pptn, Me2CO pptn, acid treatment at pH 5.7 and pptn at pH 4.5. The equilibrium constant for pyruvate + n-acetyl-D-mannosamine ++ /V-acetylneuraminidate at 37° is 0.64. The Km for A-acetylneuraminic acid is 3.9mM in phosphate at pH 7.2 and 37°. [Comb and Roseman Methods in Enzymology 5 391 1962). The enzyme from Hogg kidney (cortex) has been purified 1700 fold by extraction with H20, protamine sulphate pptn, (NH4)2S04 pptn, heat treatment between 60-80°, a second (NH4)2S04 pptn and starch gel electrophoresis. The Km for A-acetylneuraminic acid is 1.5mM. [Brunetti et al. JBC 237 2447 1962). 

Fig. 20. Proposed reaction scheme of sialate-pyruvate lyase from Clostridium perfringens. Based on data from refs. [33,892]. 

The doubly labelled sialic acids N-acetyl-[2-i4C,9-3H]neuraminic acid (NOhle and Schauer 1981) and N-glycolyl-[2-i4C,9-3H]neuraminic acid (NOhle et al. 1982) were prepared from sodium[2-i4C]pyruvate and either N-acetyl-[6-3H]mannosamine or N-glycolyl-[6- H]mannosamine, with the aid of the N-acetylneuraminate lyase from Clostridium perfringens. The metabolic fate of these compounds was studied after oral and intravenous application to mice and rats. 

A-Acetylneuraminic acid aldolase (NeuA EC 4.1.3.3) catalyzes the reversible addition of pyruvate to A-acetyl-D-mannosamine (1) to form the parent sialic acid (3) (Fig. 4). The NeuA lyases found in both bacteria and animals are type I enzymes that form an enamine intermediate with pyruvate and promote a j/-face attack to the aldehyde carbonyl group with formation of a (4S) configurated stereo center. Enzyme preparations from Clostridium perfringens and E. coli are commercially available, and the latter enzyme has been cloned, overexpressed [44,45], and its three-dimensional structure determined [46]. The enzyme has a broad pH optimum around 7.5 and is quite stable in solution at ambient temperature [47]. 

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