Bang-Ho Song

Inhibition of protein kinase CKII activity by resveratrol, a natural compound in red wine and grapes

Resveratrol is a phytoalexin found in grapes and other foods that has been shown to have anticancer and anti-inflammatory effects. Because protein kinase CKII is involved in cell proliferation and oncogenesis, we examined whether resveratrol could modulate CKII activity. Resveratrol was shown to inhibit the phosphotransferase activity of CKII with IC(50) of about 10 microM. Steady state studies revealed that resveratrol acted as a competitive inhibitor with respect to the substrate ATP. A value of 1.2 microM was obtained for the apparent K(i). Resveratrol also inhibited the catalytic reaction of CKII with GTP as substrate. Furthermore, resveratrol inhibits endogenous CKII activity on protein substrates in HeLa cell lysates. These results suggest that resveratrol is likely to function by inhibiting oncogenic disease, at least in part, through the inhibition of CKII activity.

Purification and characterization of thermostable cytidine deaminase encoded by the Bacillus caldolyticus cdd gene

Abstract A 565 bp DNA stretch coding for the Bacillus caldolyticus DSM405 cdd gene was cloned by PCR amplification and sequenced. Nucleotide sequence analysis to the cloned DNA stretch revealed an open reading frame of 396 bp encoding a deduced polypeptide of 132 amino acids with a molecular mass of 14,235 Da. Comparison of the deduced polypeptide of the B. caldolyticus cdd gene to that of the Bacillus subtilis cdd showed a 71% sequence homology. Thermostable cytidine deaminase expressed in Escherichia coli JF611/pCJH53 was purified through rapid procedures consisting of anionic-exchange column chromatography and FPLC. Finally purified enzyme showed 167.5 u/mg of specific activity, corresponding to about 94-fold purification with respect to the crude extract. Molecular mass of the monomer was estimated to be about 14-kDa by SDS-polyacrylamide gel electrophoresis (PAGE), and tetrameric configuration of the native enzyme was supposed by gel filteration. Optimal deamination occurred at 70∼75°C and pH 7.5–8.0, and the activity was inhibited by Fe 2+ , Cu 2+ , Hg 2+ , and p -chloromercuribenzoic acid ( p CMB). The enzyme was thermostable and the apparent K m and V max values for the cytidine were 2.1 × 10 −3 M and 36.4 mM min −1 mg −1 , respectively.