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Coleman, William Joseph 2015-05-14T15:21:26Z 2015-05-14T15:21:26Z 10000-01-01 1987 1987 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. Made available in DSpace on 2015-05-14T15:21:26Z (GMT). No. of bitstreams: 2 license.txt: 4848 bytes, checksum: 96035ab3f5e1c23cc7138a224ce498bd (MD5) 8721614.PDF: 5906634 bytes, checksum: 11528a5ebf1f90e9553f740a57a66020 (MD5) Previous issue date: 1987 Embargo set by: Seth Robbins for item 78895 Lift date: Forever Reason: Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs U of I Only 222 p. Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1987. http://hdl.handle.net/2142/77685 (UMI)AAI8721614 eng Chemistry, Biochemistry The Mechanism of Chloride Activation of Oxygen Evolution in Spinach Photosystem Ii text Dissertation Ph.D. Chemistry and Chemical Engineering Biochemistry University of Illinois at Urbana-Champaign