BALENDRA, S., LESIEUR, C., SMITH, T. J. and DALTON, H. (2001). Positively charged amino acids are essential for electron transfer and protein-protein interactions in the soluble methane monooxygenase complex from methylococcus capsulatus (Bath). Biochemistry, 41 (8), 2571-2579.
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The soluble methane monooxygenase (sMMO) complex from Methylococcus capsulatus (Bath) catalyses oxygen- and NAD(P)H-dependent oxygenation of methane, propene and other substrates. Whole-complex sMMO oxygenase activity requires all three sMMO components: the hydroxylase, the reductase and protein B. Also, in the presence of hydrogen peroxide, the hydroxylase alone catalyses substrate oxygenation via the peroxide shunt reaction. We investigated the effect of amine cross-linking on hydroxylase activity in order to probe the role of a gross conformational change that occurs in the hydroxylase upon binding of the other protein components. The cross-linker inhibited hydroxylase activity in the whole complex but this effect was due to covalent modification of primary amine groups rather than cross-linking. Covalent modification of arginine side-chains on the hydroxylase had a similar effect but, most remarkably, neither form of modification affected the activity of the hydroxylase via the peroxide shunt reaction. It was shown that covalent modification of positively charged groups on the hydroxylase, which occurred at multiple sites, interfered with its physical and functional interactions with protein B and with the passage of electrons from the reductase. These results indicate that protein B and the reductase of the sMMO complex interact via positively charged groups on the surface of the hydroxylase to induce a conformational change that is necessary for delivery of electrons into the active site of the hydroxylase. Modification of positively charged groups on protein B had no effect on its function, consistent with the hypothesis that positively charged groups on the hydroxylase interact with negative charges on protein B. Thus, we have discovered a means of specifically inactivating the interactions between the sMMO complex while preserving the catalytic activity of the hydroxylase active site which provides a new method of studying intercomponent interactions within sMMO.
|Additional Information:||Figures and tables reproduced here with permission from Biochemistry 41 (8), 2571-2579. Copyright 2002 American Chemical Society.|
|Research Institute, Centre or Group:||Biomedical Research Centre|
|Depositing User:||Ann Betterton|
|Date Deposited:||11 Oct 2007|
|Last Modified:||21 Dec 2010 11:30|
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