Lipid-Transfer Protein (LTP) from Wheat Kernel Possesses a Weak, Specific Esterase-like Activity Towards Short Chain Fatty Acid Esters

Summary

31P nuclear magnetic resonance studies showed that palmitoyl lysophosphatidyl-choline is slowly hydrolyzed when incubated with LTP from wheat. Kinetic measurements using paranitrophenyl esters of saturated fatty acids differing in their chain length were performed. When the fatty acid chain contained more then 12 carbons, the fatty acid was not released from the protein after cleavage of the ester bond, thus preventing new substrate binding. The thermodynamic constants for substrate binding were measured using fluorescence methods and isothermic titration calorimetry. Owing to their high affinity for the binding pocket, long chains were not released after cleavage, whereas short chain substrates were freed after hydrolysis, allowing a continuous turn-over of hydrolysis. These observations suggest that wheat LTP possesses a catalytic site susceptible of specific cleaving of ester bonds involving fatty acids. The activity is pH-dependent with a half-effect of about pH 5.4. We performed nuclear magnetic resonance titration of the 3 histidines contained in the protein, one of which (His 35) had a pKa of 5.5. Moreover, the LTP from maize, which possesses a structure closely related to that of wheat protein, had no esterase activity and contained no histidine. The activity was lost after chemical modification of the 3 histidines, suggesting that it was histidine-dependent. The in vivo significance of this weak activity is still unknown. However, this finding provides new insights into the structure-function relationship of plant lipid-binding proteins.