Serine hydroxymethyltransferase from spinach leaf mitochondria. Purification and characterization

Abstract

A mitochondrial serine hydroxymethyltransferase (EC 2.1.2.1) has for the first time been purified close to homogeneity from a photosynthetically active tissue, spinach (Spinacea oleracea L. cv Viking II) leaves. The specific activity of the enzyme was 7.8 μmol (mg protein)−1 min−1 using L‐serine as substrate. The enzyme was stable for at least 8 weeks at 4°C in the presence of folate. The pH optimum was at pH 8.5 where the enzyme had a Km for L‐serine of 0.9 mM. Carboxymethoxylamine was a strong competitive inhibitor with a K1 of 1.4 μM. An absorption spectrum taken of the enzyme in the presence of glycine and tetrahydrofolate showed a peak at 492 nm, probably originating from a substrate‐enzyme complex. The molecular weight obtained by gel filtration was 209 kDa. Sodium dodecyl sulphate‐polyacrylamide gel electrophoresis of the purified enzyme showed that the apparent molecular weight of the subunit was 53 kDa, indicating four subunits.