Identification of peptides in wheat germ hydrolysate that demonstrate calmodulin-dependent protein kinase II inhibitory activity

Hydrolysis of wheat germ by proteases resulted in bioactive peptides that demonstrated an inhibitory effect against the vasoconstrictive Ca2+-calmodulin (CaM)-dependent protein kinase II (CaMK II). The hydrolysate by thermolysin (1.0 wt%, 5 h) showed a particularly potent CaMK II inhibition. As a result of mixed mode high-performance liquid chromatography of thermolysin hydrolysate with pH elution gradient ranging between 4.8 and 8.9, the fraction eluted at pH 8.9 was the most potent CaMK II inhibitor. From this fraction, Trp-Val and Trp-Ile were identified as CaMK II inhibitors. In Sprague-Dawley rats, an enhanced aortic CaMK II activity by 1 μM phenylephrine was significantly (p < 0.05) suppressed by 15-min incubation with 300 μM Trp-Val or Trp-Ile. On the basis of Ca2+-chelating fluorescence and CaMK II activity assays, it was concluded that Trp-Val and Trp-Ile competed with Ca2+-CaM complex to bind to CaMK II with Ki values of 5.4 and 3.6 μM, respectively.