48
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: not found

      Reducing the environmental sensitivity of yellow fluorescent protein. Mechanism and applications.

      Read this article at

      ScienceOpenPublisherPubMed
      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Yellow mutants of the green fluorescent protein (YFP) are crucial constituents of genetically encoded indicators of signal transduction and fusions to monitor protein-protein interactions. However, previous YFPs show excessive pH sensitivity, chloride interference, poor photostability, or poor expression at 37 degrees C. Protein evolution in Escherichia coli has produced a new YFP named Citrine, in which the mutation Q69M confers a much lower pK(a) (5.7) than for previous YFPs, indifference to chloride, twice the photostability of previous YFPs, and much better expression at 37 degrees C and in organelles. The halide resistance is explained by a 2.2-A x-ray crystal structure of Citrine, showing that the methionine side chain fills what was once a large halide-binding cavity adjacent to the chromophore. Insertion of calmodulin within Citrine or fusion of cyan fluorescent protein, calmodulin, a calmodulin-binding peptide and Citrine has generated improved calcium indicators. These chimeras can be targeted to multiple cellular locations and have permitted the first single-cell imaging of free [Ca(2+)] in the Golgi. Citrine is superior to all previous YFPs except when pH or halide sensitivity is desired and is particularly advantageous within genetically encoded fluorescent indicators of physiological signals.

          Related collections

          Author and article information

          Journal
          J Biol Chem
          The Journal of biological chemistry
          American Society for Biochemistry & Molecular Biology (ASBMB)
          0021-9258
          0021-9258
          Aug 03 2001
          : 276
          : 31
          Affiliations
          [1 ] Howard Hughes Medical Institute, University of California, San Diego, La Jolla, California 92093-0647, USA.
          Article
          S0021-9258(20)80385-8
          10.1074/jbc.M102815200
          11387331
          e73c4f1e-65cf-4e07-8522-8c38d6db20e1
          History

          Comments

          Comment on this article