Biophysics and the Challenges of Emerging Threats

  • Joseph D. Puglisi

Table of contents

  1. Front Matter
    Pages i-vii
  2. Eric R. Henry, William A. Eaton
    Pages 1-20
  3. Timothy V. Pyrkov, Anton O. Chugunov, Nikolay A. Krylov, Dimitry E. Nolde, Roman G. Efremov
    Pages 21-41
  4. Angela M. Gronenborn
    Pages 43-50
  5. Saulius KlimaŠauskas, Zita LiutkeviČiŪtĖ, Dalia DaujotytĖ
    Pages 51-64
  6. Corey W. Liu, Viktor Y. Alekseyev, Jeffrey R. Allwardt, Alexander J. Bankovich, Barbara J. Cade-Menun, Ronald W. Davis et al.
    Pages 65-81
  7. Colin EcheverrÍa Aitken, R. Andrew Marshall, Joseph D. Pugi
    Pages 83-99
  8. Ian M. Robertson, Leo Spyracopoulos, Brian D. Sykes
    Pages 101-119
  9. Back Matter
    Pages 157-179

About these proceedings

Introduction

Single-molecule techniques eliminate ensemble averaging, thus revealing transient or rare species in heterogeneous systems [1–3]. These approaches have been employed to probe myriad biological phenomena, including protein and RNA folding [4–6], enzyme kinetics [7, 8], and even protein biosynthesis [1, 9, 10]. In particular, immobilization-based fluorescence te- niques such as total internal reflection fluorescence microscopy (TIRF-M) have recently allowed for the observation of multiple events on the millis- onds to seconds timescale [11–13]. Single-molecule fluorescence methods are challenged by the instability of single fluorophores. The organic fluorophores commonly employed in single-molecule studies of biological systems display fast photobleaching, intensity fluctuations on the millisecond timescale (blinking), or both. These phenomena limit observation time and complicate the interpretation of fl- rescence fluctuations [14, 15]. Molecular oxygen (O) modulates dye stability. Triplet O efficiently 2 2 quenches dye triplet states responsible for blinking. This results in the for- tion of singlet oxygen [16–18]. Singlet O reacts efficiently with organic dyes, 2 amino acids, and nucleobases [19, 20]. Oxidized dyes are no longer fluor- cent; oxidative damage impairs the folding and function of biomolecules. In the presence of saturating dissolved O , blinking of fluorescent dyes is sup- 2 pressed, but oxidative damage to dyes and biomolecules is rapid. Enzymatic O -scavenging systems are commonly employed to ameliorate dye instability. 2 Small molecules are often employed to suppress blinking at low O levels.

Keywords

Base Biophysics DNA Translation biochemistry biotechnology fluorescence magnetic resonance

Editors and affiliations

  • Joseph D. Puglisi
    • 1
  1. 1.SMRL & Dept. of Structural Biology D105A Fairchild Science BuildingStanford UniversityStanfordUSA

Bibliographic information

  • DOI https://doi.org/10.1007/978-90-481-2368-1
  • Copyright Information Springer Netherlands 2009
  • Publisher Name Springer, Dordrecht
  • eBook Packages Physics and Astronomy
  • Print ISBN 978-90-481-2367-4
  • Online ISBN 978-90-481-2368-1
  • Series Print ISSN 1874-6500
  • About this book