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The inhibition of a membrane-bound enzyme as a model for anaesthetic action and drug toxicity.

ABSTRACT: The inhibition of the membrane-bound enzyme cytochrome c oxidase by aliphatic n-alcohols and other neutral organic compounds was studied as a model for anaesthetic action and drug toxicity. The n-alcohols (C1 to C14) displayed a variation in inhibition constant of over 500,000-fold. The inhibition constants correlated well with the number of carbon atoms in the n-alcohols and also their n-octanol/water partition coefficients. General anaesthetic potency is known to be similarly well correlated with octanol/water partition coefficients. The free-energy change for transferring a methylene group of the n-alcohol to the more hydrophobic environment bound to the enzyme is similar to that for transferring a methylene group from water to pure alcohol. These results are consistent with the n-alcohols inhibiting by binding to an octanol-like environment on the enzyme or the protein/phospholipid interface. Neither negatively charged carboxylates nor positively charged amine analogues were observed to cause any inhibition, indicating that this postulated binding site may be uncharged. Inhibition of cytochrome c oxidase by n-alcohols was also demonstrated in both bovine heart and rat liver sonicated submitochondrial fragments.

SUBMITTER: Hasinoff BB

PROVIDER: S-EPMC1138329 | biostudies-other | 1989 Feb

REPOSITORIES: biostudies-other

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Colorimetric <i>in situ</i> assay of membrane-bound enzyme based on lipid bilayer inhibition of ion transport.

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2005-08-24 | GSE3180 | GEO

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Project description:The MAPK/ERK kinase MEK is a shared effector of the frequent cancer drivers KRAS and BRAF that has long been pursued as a drug target in oncology1, and more recently in immunotherapy2,3 and ageing4. However, many MEK inhibitors are limited owing to on-target toxicities5-7 and drug resistance8-10. Accordingly, a molecular understanding of the structure and function of MEK within physiological complexes could provide a template for the design of safer and more effective therapies. Here we report X-ray crystal structures of MEK bound to the scaffold KSR (kinase suppressor of RAS) with various MEK inhibitors, including the clinical drug trametinib. The structures reveal an unexpected mode of binding in which trametinib directly engages KSR at the MEK interface. In the bound complex, KSR remodels the prototypical allosteric pocket of the MEK inhibitor, thereby affecting binding and kinetics, including the drug-residence time. Moreover, trametinib binds KSR-MEK but disrupts the related RAF-MEK complex through a mechanism that exploits evolutionarily conserved interface residues that distinguish these sub-complexes. On the basis of these insights, we created trametiglue, which limits adaptive resistance to MEK inhibition by enhancing interfacial binding. Our results reveal the plasticity of an interface pocket within MEK sub-complexes and have implications for the design of next-generation drugs that target the RAS pathway.

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Assessment of Awareness of Local Anaesthetic Systemic Toxicity (LAST) among Postgraduate Trainees

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