1. Green Fluorescent Protein
a B/MB senior seminar
brought to you by Colm O’Carroll

4. This presentation will cover
The structural aspects of GFP which make fluorescence possible
The advantages of using GFP and GFP mutants over other fluorescent markers
The use of GFP to monitor viral movement in plants

5. The Green Fluorescent Protein

6. GFP’s unique structure
Composed of 238 amino acids
“Paint in a can”
Each monomer composed of a central -helix surrounded by an eleven stranded cylinder of anti-parallel -sheets
Cylinder has a diameter of about 30A and is about 40A long
Fluorophore located on central helix

7. The Active Site

9. The Fluoropore Active Site
Ser65-Tyr66-Gly67
Deprotonated phenolate of Tyr66 is cause of fluorescence
Forster Cycle (1949-Theodor Forster)
Proton transfer to His148

10. Fluorophore formation
One limitation of wtGFP is its slow rate of fluorescence acquisition in vivo
Renaturation most likely by a parallel pathway
Oxidation of Fluoropore (2-4 hours)
Two step process