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Open Access Publications from the University of California

Scholarly Works (2 results)

  • Article
  • Peer Reviewed
UCLA Previously Published Works (2014)
Compared to thylakoid and inner membrane proteins in cyanobacteria, no structure-function information is available presently for integral outer-membrane proteins (OMPs). The Slr1270 protein from the cyanobacterium Synechocystis 6803, over-expressed in Escherichia coli, was refolded, and characterized for molecular size, secondary structure, and ion-channel function. Refolded Slr1270 displays a single band in native-electrophoresis, has an α-helical content of 50-60%, as in E. coli TolC with which it has significant secondary-structure similarity, and an ion-channel function with a single-channel conductance of 80-200pS, and a monovalent ion (K(+):Cl(-)) selectivity of 4.7:1. The pH-dependence of channel conductance implies a role for carboxylate residues in channel gating, analogous to that in TolC.
    Cover page: Structure–function of cyanobacterial outer‐membrane protein, Slr1270: Homolog of Escherichia coli drug export/colicin import protein, TolC
    • Article
    • Peer Reviewed

    Trans-membrane Signaling in Photosynthetic State Transitions REDOX- AND STRUCTURE-DEPENDENT INTERACTION IN VITRO BETWEEN STT7 KINASE AND THE CYTOCHROME b6f COMPLEX* * This work was supported by National Institutes of Health Grant GMS-038323 (to W. A. C.), Purdue University Center for Cancer Research Grant P30 CA23168 (to W. A. C.), and University of California, San Diego/Los Angeles NIDDK, National Institutes of Health Diabetes Research Center Grant P30 DK063491 (to J. P. W.). A preliminary account was presented at the meeting of the Biophysical Society, February 27-March 2, 2016, Los Angeles, CA ((2016) Biophys. J. 110, 91a). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

    UCLA Previously Published Works (2016)
    Trans-membrane signaling involving a serine/threonine kinase (Stt7 in Chlamydomonas reinhardtii) directs light energy distribution between the two photosystems of oxygenic photosynthesis. Oxidation of plastoquinol mediated by the cytochrome b6f complex on the electrochemically positive side of the thylakoid membrane activates the kinase domain of Stt7 on the trans (negative) side, leading to phosphorylation and redistribution ("state transition") of the light-harvesting chlorophyll proteins between the two photosystems. The molecular description of the Stt7 kinase and its interaction with the cytochrome b6f complex are unknown or unclear. In this study, Stt7 kinase has been cloned, expressed, and purified in a heterologous host. Stt7 kinase is shown to be active in vitro in the presence of reductant and purified as a tetramer, as determined by analytical ultracentrifugation, electron microscopy, and electrospray ionization mass spectrometry, with a molecular weight of 332 kDa, consisting of an 83.41-kDa monomer. Far-UV circular dichroism spectra show Stt7 to be mostly α-helical and document a physical interaction with the b6f complex through increased thermal stability of Stt7 secondary structure. The activity of wild-type Stt7 and its Cys-Ser mutant at positions 68 and 73 in the presence of a reductant suggest that the enzyme does not require a disulfide bridge for its activity as suggested elsewhere. Kinase activation in vivo could result from direct interaction between Stt7 and the b6f complex or long-range reduction of Stt7 by superoxide, known to be generated in the b6f complex by quinol oxidation.
      Cover page: Trans-membrane Signaling in Photosynthetic State Transitions REDOX- AND STRUCTURE-DEPENDENT INTERACTION IN VITRO BETWEEN STT7 KINASE AND THE CYTOCHROME b6f COMPLEX* * This work was supported by National Institutes of Health Grant GMS-038323 (to W. A. C.), Purdue University Center for Cancer Research Grant P30 CA23168 (to W. A. C.), and University of California, San Diego/Los Angeles NIDDK, National Institutes of Health Diabetes Research Center Grant P30 DK063491 (to J. P. W.). A preliminary account was presented at the meeting of the Biophysical Society, February 27-March 2, 2016, Los Angeles, CA ((2016) Biophys. J. 110, 91a). The authors declare that they have no conflicts of interest with the contents of this article. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
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