The Nature of the Hydrophobic n-Alkanol Binding Site within the C1 Domains of Protein Kinase C
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- 作者：Simon J. Slater ; Steve A. Malinowski ; and Christopher D. Stubbs
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：June 15, 2004
- 年：2004
- 卷：43
- 期：23
- 页码：7601 - 7609
- 全文大小：140K
- 年卷期：v.43,no.23(June 15, 2004)
- ISSN：1520-4995
文摘
The activator-binding sites within the C1 domains of protein kinase C (PKC) are also able tobind alcohols and anesthetics. In this study, the nature of the interaction of these agents with the hydrophobicregion within the C1 domains was investigated and a structure-activity relationship for the alcohol effectswas obtained. The effects of a series of n-alkanols on PKC
activity, determined using an in vitro assaysystem that lacked lipids, were found to be a nonlinear function of the chain length. In the absence ofphorbol ester or diacylglycerol, 1-octanol potently activated PKC in a concentration-dependent manner,while 1-heptanol was completely without effect, despite differing by one methylene unit. The minimalstructural requirement for the activating effect corresponded to R-CH(OH)-(CH2)n-CH3, where R = Hor an alkyl group and n 
6. Consistent with this, 2-octanol, for which n = 5, was without effect on theactivity, even though this alcohol is only marginally less hydrophobic than 1-octanol, whereas 2-nonanol,for which n = 6, was able to produce activity. Importantly, it was found that PKC was activated to agreater extent by R-2-nonanol than by the S enantiomer. The potentiation of phorbol ester-induced,membrane-associated PKC activity by long-chain n-alkanols reported previously (Slater, S. J., Kelly,M. B., Larkin, J. D., Ho, C, Mazurek, A, Taddeo, F. J., Yeager, M. D., Stubbs, C. D. (1997) J. Biol.Chem. 272, 6167-6173), was also found here for nonmembrane associated PKC, indicating that thiseffect is an intrinsic property of the enzyme rather than a result of membrane perturbation. Overall, theresults suggest that the alcohol-binding sites within the C1 domains of PKC contain spatially distincthydrophilic and hydrophobic regions that impose a high degree of structural specificity on the interactionsof alcohols and other anesthetic compounds, as well as diacylglycerols and phorbol esters.
activity, determined using an in vitro assaysystem that lacked lipids, were found to be a nonlinear function of the chain length. In the absence ofphorbol ester or diacylglycerol, 1-octanol potently activated PKC in a concentration-dependent manner,while 1-heptanol was completely without effect, despite differing by one methylene unit. The minimalstructural requirement for the activating effect corresponded to R-CH(OH)-(CH2)n-CH3, where R = Hor an alkyl group and n 6. Consistent with this, 2-octanol, for which n = 5, was without effect on theactivity, even though this alcohol is only marginally less hydrophobic than 1-octanol, whereas 2-nonanol,for which n = 6, was able to produce activity. Importantly, it was found that PKC was activated to agreater extent by R-2-nonanol than by the S enantiomer. The potentiation of phorbol ester-induced,membrane-associated PKC activity by long-chain n-alkanols reported previously (Slater, S. J., Kelly,M. B., Larkin, J. D., Ho, C, Mazurek, A, Taddeo, F. J., Yeager, M. D., Stubbs, C. D. (1997) J. Biol.Chem. 272, 6167-6173), was also found here for nonmembrane associated PKC, indicating that thiseffect is an intrinsic property of the enzyme rather than a result of membrane perturbation. Overall, theresults suggest that the alcohol-binding sites within the C1 domains of PKC contain spatially distincthydrophilic and hydrophobic regions that impose a high degree of structural specificity on the interactionsof alcohols and other anesthetic compounds, as well as diacylglycerols and phorbol esters.