Melvin P. Klein

Paramagnetic Resonance of Fe3+ in Polycrystalline Ferrichrome A

Polycrystalline samples of iron‐containing ferrichrome A, a cyclic hexapeptide obtained from the fungus Ustillago sphaerogena, have been investigated by paramagnetic resonance. Spectra were obtained at several temperatures between 300° and 1°K; a prominent line of 400‐Oe width located at g=4.3 was observed at all temperatures, while at 1°K additional resonances at g values of 9.6, 1.3, and 1.0 were observed. The spectra are interpreted by assuming a spin Hamiltonian containing crystal‐field terms large compared with the Zeeman splittings; the crystal‐field situation is intermediate between the case of axial symmetry, with H=D[Sz2−(1/3)S(S+1)]+gβS·H and a model proposed by Castner, Newell, Holton, and Slichter to explain certain iron resonances occurring at g=4.3, with H=E(Sx2−Sy2)+gβS·H. We have computed g values, energy eigenvalues, and eigenfunctions to be expected for the region between these two extremes, and the results should be useful in interpreting similar spectra due to iron situated in strong c...

CHEMICAL EFFECTS ON CORE ELECTRON BINDING ENERGIES IN IODINE AND EUROPIUM

A theoretical and experimental study was made of the shift in atomic core‐electron binding energies caused by the chemical environment. Two models are presented to account for these “chemical shifts.” The first uses an energy cycle to break the core‐electron binding energies into a free‐ion contribution and a classical Madelung energy contribution. The Madelung energy contributes a significant part of the binding‐energy shift. It can, in principle, be evaluated rigorously although there is some ambiguity as to a surface correction. The reference level for binding energies must also be considered in comparing theory with experiment (or in comparing experimental shifts with one another). Electronic relaxation could also introduce errors of ∼1 eV in shift measurements. The second, more approximate, model consists of a “charged‐shell” approximation for bonding electrons in atomic complexes. It gives semiquantitative estimates of shifts and demonstrates the relationship between bond polarity and core‐electron ...

The state of manganese in the photosynthetic apparatus. 3. Light-induced changes in X-ray absorption (K-edge) energies of manganese in photosynthetic membranes

Abstract Photosynthetic water oxidation by higher plants proceeds as though five intermediates, S0-S4, operate in a cyclic fashion. In this study of the manganese involvement in the process, a low temperature EPR signal is used as an indicator of S-state composition for manganese X-ray absorption K-edge measurements of a spinach Photosystem II preparation. A dramatic change is observed in the edge properties between samples prepared in states S1 and either S2 or S3, establishing a direct relation between the local environment of Mn and the S-state composition. Samples in S2 or S3 exhibit a broadening of the principal absorption peak and a shift to higher energy by as much as 2.5 eV relative to S1 samples. The magnitude of these changes is directly related to the EPR signal intensity induced by illumination. Models are discussed in which these data may be interpreted in terms of a conformation-induced change in Mn ligation and/or oxidation during the S1 to S2 transition.

The state of manganese in the photosynthetic apparatus: 4. Structure of the manganese complex in Photosystem II studied using EXAFS spectroscopy. The S1 state of the O2-evolving Photosystem II complex from spinach

Abstract The structure of the Mn complex in the oxygen-evolving system and its mechanistic relation to photosynthetic oxygen evolution are poorly understood, though many studies have established that membrane-bound Mn plays an active role. Recently established procedures for isolating oxygen-evolving subchloroplast Photosystem II (PS II) preparations and the discovery of a light-induced multiline EPR signal attributable to the S2 state of the O2-evolving complex have facilitated the preparation of samples well characterized in the S1 and S2 states. We have used extended X-ray absorption fine structure (EXAFS) spectroscopy to probe the ligand environment of Mn in PS II particles from spinach, and in this report we present our results. The essential feature of the EXAFS results are that at least two Mn atoms per PS II reaction center occur as a binuclear species with a metal-metal distance of approx. 2.7 A, with low Z atoms, N or O, at a distance of approx. 1.75 A and at approx. 1.98 A, which are characteristic of bridging and terminal ligands. These results agree well with those derived from whole chloroplasts that provided the first evidence for a binuclear manganese complex (Kirby, J.A., Robertson, A.S., Smith, J.P., Thompson, A.C., Cooper, S.R. and Klein, M.P. (1981) J. Am. Chem. Soc. 103, 5529–5537).

Solution conformation of ferrichrome, a microbial iron transport cyclohexapeptide, as deduced by high resolution proton magnetic resonance☆

Abstract The proton magnetic resonance spectra of the cyclohexapeptides deferriferrichrome and its Al 3+ chelate (alumichrome) dissolved in water and in dimethyl sulfoxide are compared. The spin-spin coupling constants ( J NC ) between the amide NH and C α H protons suggest that the metal affects the conformation of the δ- N -acetyl-δ- N -hydroxy- l -ornithine sidechains more than that of the peptide backbone. The data are consistent with a structure containing two transannular H-bonds in both deferriferrichrome and alumichrome as in the Schwyzer model of cyclohexapeptides. Metal chelation induces chemical shifts which are pronounced for all of the amide NHs and markedly reduces the solvent interactions with four of the NHs. The solution conformation of the chelate is in good agreement with the X-ray structure of crystalline ferrichrome A, a closely related compound.

The state of manganese in the photosynthetic apparatus: 5. The chloride effect in photosynthetic oxygen evolution. Is halide coordinated to the EPR-active manganese in the O2-evolving complex? Studies of the substructure of the low-temperature multiline EPR signal

Abstract The role of chloride in the manganese-containing oxygen-evolving complex of Photosystem II has been studied by observing the amplitude of the multiline EPR signal as a function of Cl − concentration or when Cl − is replaced by Br − or F − . The correlation of the multiline EPR signal intensity and O 2 activity with the concentration of Cl − shows that chloride is involved in oxygen evolution at the S 2 or earlier S states, and that it is necessary for the production of an EPR-detectable S 2 state. We have developed a new method for the preparation of subchloroplast PS II particles containing Br − and F − ) and have used these particles for studying the EPR fine structure at high resolution. The fine structure shows a multiplet of 4–6 lines with 10–15 G spacing; at the resolution of our experiment there are no significant differences between the Cl − -and Br − -containing samples, suggesting that the halide is not a ligand of the EPR-active Mn. Various structural possibilities for the Mn complex, which would account for the observed fine structure of the multiline EPR spectrum are discussed.

Assignment of the g = 4.1 ERP signal to manganese in the S2 state of the photosynthetic oxygen-evolving complex: An X-ray absorption edge spectroscopy study☆

X-ray absorption spectroscopy at the Mn K-edge has been utilized to study the origin of the g = 4.1 EPR signal associated with the Mn-containing photosynthetic O2-evolving complex. Formation of the g = 4.1 signal by illumination of Photosystem II preparations at 140 K is associated with a shift of the Mn edge inflection point to higher energy. This shift is similar to that observed upon formation of the S2 multiline EPR signal by 190 K illumination. The g = 4.1 signal is assigned to the Mn complex in the S2 state.

A versatile loop-gap resonator probe for low-temperature electron spin-echo studies

We have recently constructed a new high-power pulsed EPR spectrometer for use in biological electron spin-echo (ESE) studies. This spectrometer produces 25 ns microwave pulses with power levels up to one kilowatt. The bandwidth requirement for admitting such narrow pulses limits the quality factor, Q, for the EPR sample cavity or resonator to a value of a few hundred. A conventional microwave cavity, such as the TElo2 cavity often used in EPR, has a critically coupled Q of several thousand. Thus the Q must be greatly reduced to meet the bandwidth requirements for pulsed EPR. Without the asset of very high Q, the large volume and resultant poor filling factor of such a cavity limit sensitivity and increase the microwave power necessary to create a given microwave magnetic field amplitude. The stripline transmission cavity of Mims (I) provides a well-tested alternative for pulsed EPR. This device has a high filling factor and appropriately low Q, but is inconvenient to use because samples are placed directly into the cavity rather than into conventional EPR tubes, precluding measurements of pulsed and conventional EPR with identical samples. In recent years, other reduced volume resonators have been introduced to EPR (2-4). We have constructed a low-temperature pulsed EPR probe built around the loopgap resonator of Froncisz and Hyde (2), taking advantage of its moderate Q and high filling factor. This loop-gap probe mounts in a liquid He immersion dewar. The immersion dewar provides economical operation and excellent stability at temperatures as low as 1.5 K. The first loop-gap probe we constructed used a movable inductive loop to variably couple microwave power to the resonator. This system suffered from excessive microphonics, as the coupling loop vibrated in the bubbling liquid He. This led us to consider a system in which microwave power is coupled to the loop-gap resonator directly through a waveguide or cavity. In designing such a system we were constrained by the relatively small i.d. of the liquid helium dewar (3.2 cm). Many of our samples are poised in an EPR-active state at cryogenic temperatures and are unstable at room temperature. Thus an additional constraint was provided by the necessity of introducing these samples directly into the resonator while the probe system was immersed in liquid He. This capability would additionally allow us to change samples without removing the entire probe structure from the liquid He, conserving both liquid He and experimental time. We also planned to use the design for conventional EPR and for ENDOR experiments. In this note we present details of

Profile of a focussed collimated laser beam near the focal minimum characterized by fluorescence correlation spectroscopy

Central to the application of fluorescence correlation spectroscopy, to measure the self‐diffusion coefficients and average concentration of fluorescent molecules in a volume determined by a focussed laser beam, is the determination of the focal spot size. As the focal spot size in the sample plane is varied by displacing either the focusing lens or sample position along the beam axis, the diffusion time and average number of molecules vary in a parabolic manner. Analysis of the parameters of the parabola leads to estimates of the beam radius at the waist. The results agree with theoretical predictions and provide an independent measurement of the beam profile.

Perspectives on the structure of the photosynthetic oxygen evolving manganese complex and its relation to the Kok cycle.

This review describes the progress in our understanding of the structure of the Mn complex in Photosystem II over the last two decades. Emphasis is on the research from our laboratory, especially the results from X-ray absorption spectroscopy, low temperature electron paramagnetic resonance and electron spin echo envelope modulation studies. The importance of the interplay between electron paramagnetic resonance studies and X-ray absorption studies, which has led to a description of the oxidation states of manganese as the enzyme cycles through the Kok cycle, is described. Finally, the path, by which our group has utilized these two important methods to arrive at a working structural model for the manganese complex that catalyzes the oxidation of water to dioxygen in higher plants and cyanobacteria, is explained.