Purification of 5-aminolaevulinate synthase from liver mitochondria of chick embryo

Abstract

5-Aminolaevulinate synthase from chick-embryo liver mitochondria has, for the first time, been purified to homogeneity in its native non-degraded form by molecular sieve chromatography, chromatofocusing and affinity chromatography. The enzyme has a minimum molecular weight of 68000 as determined by sodium dodecylsulphate/polyacrylamide gel electrophoresis and a specific activity of 35000 units/mg of protein. This result conflicts with the previous report of Whiting, M.J. and Granick, G. [(1976) J. Biol. Chem. 251, 1340-1346] that the chick embryo enzyme has a molecular weight of 49000. We show here that the purified form can be degraded proteolytically to a smaller form of molecular weight around 50000 while retaining full enzymatic activity. It seem evident, therefore, that the enzyme isolated by Whiting & Granick (1976) was degraded. We have further established by pulse-labelling studies and immunoprecipitation that the enzyme isolated by our new and rapid procedure has the same minimum molecular weight as that which exists in vivo.