PURIFICATION AND PROPERTIES OF DIAMINOPIMELATE DECARBOXYLASE FROM ESCHERICHIA COLI

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Abstract
Diaminopimelate decarboxylase from a soluble extract of E. coli A.T.C.C. 9637 was purified 200-fold. The purified enzyme showed only 1 component in the ultracentrifuge, with a sedimentation coefficient of 5-4s. The major peak and 3 smaller peaks were observed on electrophoresis of the enzyme at pH 8-9. The molecular wt. of the enzyme was approximately 2000 000. The catalytic constant was 2000mol. of mesodiaminopimelic acid decomposed/min./mol. of enzyme, at 37[degree]. The relative rates of decarboxylation at 25[degree], 37[degree] and 45[degree] were 0.171-0:1-6. The Michaelis constant was 1.7 m[image] and the optimum pH was 6.7[long dash]6.8 at 37[degree]. There was an excess of acidic amino-acids over basic amino-acids in the enzyme, which was found only on basic cellulose derivatives at pH 6-8. The enzyme had an absolute requirement for pyridoxal phosphate as a cofactor; no other derivative of pyridoxine had activity. A thiol compound (of which 2,3-dimercaptopropan-1-o1 was the most effective) was also needed as an activator. In the presence of 2,3-dimercaptopropan-1-ol (1 m[image]), heavy-metal ions (Cu2+, Hg2+) did not inhibit the enzyme, but there was inhibition by several amino-acids with analogous structures to diaminopimelate (generally at high concentrations relative to the substrate). Penicillamine was inhibitory at relatively low concentrations; its action was prevented by pyridoxal phosphate.