The Mechanism of Chloride Activation of Oxygen Evolution in Spinach Photosystem Ii
Coleman, William Joseph
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Permalink
https://hdl.handle.net/2142/77685
Description
Title
The Mechanism of Chloride Activation of Oxygen Evolution in Spinach Photosystem Ii
Author(s)
Coleman, William Joseph
Issue Date
1987
Department of Study
Chemistry and Chemical Engineering
Discipline
Biochemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Chemistry, Biochemistry
Language
eng
Abstract
The mechanism by which ${\rm Cl}\sp-$ activates the oxygen-evolving complex (OEC) of Photosystem II (PS II) was studied by $\sp{35}{\rm Cl}$-NMR and steady-state measurements of oxygen evolution. This study represents the first attempt to directly monitor ${\rm Cl}\sp-$ binding in spinach. At low light intensity, the ${\rm Cl}\sp-$ activation curve (Hill activity vs. (${\rm Cl}\sp-$)) shows three intermediary plateaus in the concentration range 0.1-10 mM ${\rm Cl}\sp-,$ indicating kinetic cooperativity with respect to ${\rm Cl}\sp-.$ The $\sp{35}{\rm Cl}$-NMR binding curve (excess linewidth vs. (${\rm Cl}\sp-$)) for ${\rm Cl}\sp-$ depleted thylakoids and PS II membranes is not a smoothly descending hyperbola. Instead, this curve shows four sharp increases in linewidth (linewidth maxima) in the concentration range 0.1-10 mM ${\rm Cl}\sp-.$ The presence of these linewidth maxima indicates that ${\rm Cl}\sp-$ addition exposes four ${\rm Cl}\sp-$ binding sites that were not previously accessible to exchange. The pH-dependence for the excess linewidth at 0.75 mM ${\rm Cl}\sp-$ shows a maximum at pH 6.0 and two smaller maxima at pH 5.4 and 6.5. Mild heating eliminates both the linewidth maxima and the plateaus in the ${\rm Cl}\sp-$ activation curve. Hydroxylamine treatment (1.5 mM) has little effect on ${\rm Cl}\sp-$ binding, indicating that Mn is not involved. Tris-washing eliminates nearly all of the high-affinity ${\rm Cl}\sp-$ binding. Measurements of PS II membranes washed with 1.0 M NaCl or ${\rm CaCl}\sb2$ indicate that the native ${\rm Cl}\sp-$ binding mechanism requires the presence of the 33 kD extrinsic polypeptide, but that the function of the 18 kD and 24 kD extrinsic polypeptides can be replaced by 2.0 mM ${\rm Ca}\sp{2+}.$ Chloride binding is still observed after removal of all three extrinsic polypeptides, but the $\sp{35}{\rm Cl}$-NMR binding curve shows only a single, broad linewidth maximum at about 0.5 mM ${\rm Cl}\sp-.$ A hypothetical model is proposed to explain these results. This model involves ${\rm Cl}\sp-$ binding at two types of sites: (1) an intrinsic site (composed of three histidines) on the D1/D2 proteins, and (2) a set of four extrinsic sites (composed of lysines and arginines) on the 33 kD polypeptide. Possible amino acid ligands for the tetra-nuclear Mn cluster (on D1/D2) are also described.
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