Molecular characterisation of recombinant green fluorescent protein by fluorescence correlation microscopy.
The cDNA for the green fluorescent protein (GFP) of Aequorea victoria has been expressed in transformed cells of Saccharomyces cerevisiae and the recombinant GFP isolated. Protonation and deprotonation of the cloned and purified GFP produced major effects on its spectral absorption characteristics with an increase in pH enhancing the fluorescence emission of the GFP more than twofold. Finally, molecular characterisation of GFP by fluorescence correlation microscopy in a minimal target volume of 1 fL yielded a translational diffusion coefficient (DT) of 8.7 x 10(-7) cm2.sec-1, equivalent to a Stokes radius of 2.82nm for a monodisperse globular protein of 27kDa.
Terry
B R
BR
BioImage:S, Soeborg, Denmark.
Matthews
E K
EK
Haseloff
J
J
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008856:Microscopy, Fluorescence / Q000295:instrumentation; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics; D037401:Scyphozoa / Q000737:chemistry / Q000235:genetics
D018076:DNA, Complementary; D008164:Luminescent Proteins; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins