ISOLATION AND PROPERTIES OF PORCINE THYROID FUCOKINASE

A 23 000-fold purification of porcine fucokinase (ATP:6-deoxy-l-galactose 1-phosphotransferase, EC 2.7.1.52) has been achieved using a combination of ion-exchange, hydrophobic ligand, affinity, hydroxyapatite and molecular sieve chromatography. The enzyme was determined to have a subunit molecular weight of 78 180 ± 4260 by sodium dodecyl sulfate chromatography and a tetrameric molecular weight of 309 200 ± 4100 in the active state as determined by molecular sieve chromatography. The enzyme exhibits a single pH optimum at a pH value of 6.5 and gives evidence of a high order of specificity for l-fucose and ATP. The enzyme requires a divalent metal ion and this need is best satisfied by Mg2+. The activity of the enzyme is modified by a number of nucleotides. ADP is an enzyme inhibitor competitive with ATP. GDP-β-l-fucose is also an inhibitor and appears to compete with l-fucose. GDP-α-d-mannose stimulates the enzyme. A possible role for the actions of these nucleotide sugars is discussed. © 1979.