Purification and characterization of a basic lysophospholipase in germinating barley.
Lysophospholipase from germinating barley seeds has been isolated using methods which take advantage of the fact that the activity is basic, lipophilic, and contains carbohydrate. There appears to be at least three enzymatic forms of the activity, two with molecular weights at 40,000 and one at 41,000. They comigrate with a pI of 8.8 on isoelectric focusing and they all undergo deglycosylation to give a polypeptide with molecular weight 36,000, indicating 10 to 12% carbohydrate in the original glycoproteins. The enzyme is inactivated by sulfhydryl reagents and has a tendency to aggregate. The latter property may be attenuated with mercaptoethanol with which the activity is stable for more than 3 months at 4 degrees C. The most active barley enzyme has a Km of 30 microM for lysophosphatidylcholine and a Vmax, 200 mumol/min/mg. The specific activity is 20 times greater than that for lysophospholipases isolated from animal sources. It has no phospholipase, lipase, or transacylase activity. It is most active on lysophosphatidylcholine with a saturated 16 carbon or unsaturated 18 carbon chain; these are the predominant molecular species of lysophospholipid present as inclusion complexes in barley starch. The role of the barley lysophospholipases in barley germination is discussed.
D000596:Amino Acids / Q000032:analysis; D002845:Chromatography / Q000379:methods; D002523:Edible Grain / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D006023:Glycoproteins / Q000032:analysis; D001467:Hordeum / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007700:Kinetics; D008245:Lysophospholipase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D010740:Phospholipases / Q000302:isolation & purification*; D012639:Seeds; D014508:Urea
D000596:Amino Acids; D006023:Glycoproteins; D014508:Urea; D010740:Phospholipases; D008245:Lysophospholipase