Mitochondrial, plastid, and cytosolic isozymes of hexokinase from developing endosperm of Ricinus communis.
Ion-filtration chromatography on DEAE-Sephadex A-25 at pH 5.5 resolved the total hexokinase activity in a 27,000g supernatant prepared from developing castor oil (Ricinus communis L., cv. Baker 296) seeds into three distinct peaks. These were designated one, two, and three in order of elution from the column. Ion-filtration chromatography of various subcellular fractions demonstrated that isozyme one is associated with the mitochondria, isozyme two with the leucoplasts, and isozyme three is located in the cytosol. Organelle-subfractionation studies showed that hexokinase one is associated with the mitochondrial outer membrane, and hexokinase two is located in the plastid stroma. Isozyme one could be specifically solubilized from the mitochondrial outer membrane by incubation with nucleoside triphosphates or hexose monophosphates. In addition to differences in net charge and localization, the three hexokinase isozymes could be distinguished by pH optima, substrate specificity, and response to sulfhydryl-directed inhibitors. Mr values, determined by gel permeation using Sephacryl S-200, were identical for all three isozymes at 38,000. The preferred divalent metal ion for each of the hexokinases is Mg+2, and none was affected by Na+, K+, or NH+4 at their pH optima, nor by Al+3 at pH 6.6 or 7.8.
D003600:Cytosol / Q000201:enzymology; D006593:Hexokinase / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008928:Mitochondria / Q000201:enzymology*; D008970:Molecular Weight; D010944:Plants / Q000201:enzymology*; D010947:Plants, Toxic; D012278:Ricinus / Q000201:enzymology; D013045:Species Specificity; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity
D007527:Isoenzymes; D006593:Hexokinase