Characterization Of Inhibitory Effects Of NH2OH And Its N-methyl ...

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Abstract

Inorganic cofactors (Mn, Ca(2+) and Cl(-)) are essential for oxidation of H2O to O2 by Photosystem II. The Mn reductants NH2OH and its N-methyl derivatives have been employed as probes to further examine the interactions between these species and Mn at the active site of H2O oxidation. Results of these studies show that the size of a hydroxylamine derivative regulates its ability to inactivate O2 evolution activity, and that this size-dependent inhibition behavior arises from the protein structure of Photosystem II. A set of anions (Cl(-), F(-) and SO4 (2-)) is able to slow NH2OH and CH3NHOH inactivation of intact Photosystem II membranes by exerting a stabilizing influence on the extrinsic 23 and 17 kDa polypeptides. In contrast to this non-specific anion effect, only Cl(-) is capable of attenuating CH3NHOH and (CH3)2NOH inhibition in salt-washed preparations lacking the 23 and 17 kDa polypeptides. However, Cl(-) fails to protect against NH2OH inhibition in salt-washed membranes. These results indicate that the attack by NH2OH and its N-methyl derivatives on Mn occurs at different sites in the O2-evolving complex. The small reductant NH2OH acts at a Cl(-)-insensitive site whereas the inhibitions by CH3NHOH and (CH3)2NOH involve a site that is Cl(-) sensitive. These findings are consistent with earlier studies showing that the size of primary amines controls the Cl(-) sensitivity of their binding to Mn in the O2-evolving complex.

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