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-beta-L-fucose is also an inhibitor and appears to compete with L-fucose. GDP-alpha-D-mannose stimulates the enzyme. A possible role for the actions of these nucleotide sugars is discussed.
D000818:Animals; D005643:Fucose; D006156:Guanosine Diphosphate Sugars / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008722:Methods; D008970:Molecular Weight; D009711:Nucleotides / Q000494:pharmacology; D010770:Phosphotransferases / Q000302:isolation & purification*; D017853:Phosphotransferases (Alcohol Group Acceptor); D013379:Substrate Specificity; D013552:Swine; D013961:Thyroid Gland / Q000201:enzymology*
D006156:Guanosine Diphosphate Sugars; D009711:Nucleotides; D005643:Fucose; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C020153:fucokinase