Corn phosphoglycolate phosphatase: purification and properties.

Phosphoglycolate phosphatase (EC 3.1.3.18), isolated from maize leaf bundle sheath, was purified 200-fold to a specific activity of about 99 μmol mg(-1) protein · min(-1). The purification procedure included Sephadex G-75 filtration, and diethylaminoethyl-cellulose and Phospho-Ultrogel A6R chromatography. This partially purified enzyme exhibited optimum activity over a broad pH range, from pH 6.3 to pH 8.0. It displayed a very high degree of specificity for phosphoglycolate and required a divalent cation to be active; Mg(2+) was the most effective activator. Saturation curves of the Michaelis-Menten type were observed both with phosphoglycolate (Km=0.57 mmol·l(-1)) and with Mg(2+) (Km=0.015 mmol·l(-1)). The activation constant for Mg(2+) was unchanged when the pH was raised from 7.0 to 8.0. These results indicate that variations of stromal pH and Mg(2+) during the darklight transition could not directly modifity the activity of the phosphoglycolate phosphatase in maize bundle-sheath chloroplasts. The undissociated protein showed a pI of 4.95, as determined by isoelectric focusing. For the native phosphatase a molecular mass of about 61 500 Da was estimated by polyacrylamide gradient gel electrophoresis. The subunit was found to have a relative molecular mass of 31 500 Da by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is concluded that maize phosphoglycolate phosphatase is a dimer.