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PubMed2004

Nanometer-localized multiple single-molecule fluorescence microscopy.

Fitting the image of a single molecule to the point spread function of an optical system greatly improves the precision with which single molecules can be located. Centroid localization with nanometer precision has been achieved when a sufficient number of photons are collected. However, if multiple single molecules reside within a diffraction-limited spot, this localization approach does not work. This paper demonstrates nanometer-localized multiple single-molecule (NALMS) fluorescence microscopy by using both centroid localization and photobleaching of the single fluorophores. Short duplex DNA strands are used as nanoscale "rulers" to validate the NALMS microscopy approach. Nanometer accuracy is demonstrated for two to five single molecules within a diffraction-limited area. NALMS microscopy will greatly facilitate single-molecule study of biological systems because it covers the gap between fluorescence resonance energy transfer-based (<10 nm) and diffraction-limited microscopy (>100 nm) measurements of the distance between two fluorophores. Application of NALMS microscopy to DNA mapping with <10-nm (i.e., 30-base) resolution is demonstrated.

DNA↗

PubMed2002

Improvement of photosynthetic CO2 fixation at high light intensity through reduction of chlorophyll antenna size.

At elevated light intensities (greater than approximately 200 microE/[m2 x s]), the kinetic imbalance between the rate of photon excitation and thermally activated electron transport results in saturation of the rate of photosynthesis. Since maximum terrestrial solar radiation can reach 200 microE/(m2 x s), a significant opportunity exists to improve photosynthetic efficiency at elevated light intensities by achieving a kinetic balance between photon excitation and electron transport, especially in designed large-scale photosynthetic reactors in which a low-cost and efficient biomass production system is desired. One such strategy is a reduction in chlorophyll (chl) antenna size in relation to the reaction center that it serves. In this article, we report recent progress in this area of research. Light-saturation studies for CO2 fixation were performed on an antenna-deficient mutant of Chlamydomonas (DS521) and the wild type (DES15) with 700 ppm of CO2 in air. The light-saturated rate for CO2 assimilation in the mutant DS521 was about two times higher (187 micromol/[h x mg of chl]) than that of the wild type, DES15 (95 micromol/[h x mg of chl]). Significantly, a partial linearization of the light-saturation curve was also observed. These results confirmed that DS521 has a smaller relative chl antenna size and demonstrated that reduction of relative antenna size can improve the overall efficiency of photon utilization at higher light intensities. The antenna-deficient mutant DS521 can provide significant resistance to photoinhibition, in addition to improvement in the overall efficiency of CO2 fixation at high light. The experimental data reported herein support the idea that reduction in chl antenna size could have significant implications for both fundamental understanding of photosynthesis and potential application to improve photosynthetic CO2 fixation efficiency.

Animals↗