Principle Microbiology

GFP-Photo reaction:

· The GFP absorps mainly at 396 nm, followed by loss of proton in excited state to form phenolate.

· The main emmision occurs at 508 nm. Small amount of GFP absorps at 475 nm and emits green light at 508 nm.

· The GFP as well as green fluoroscent gene can be modified. The gene can be modified by mutation.

· The fluoroscent may be partially or completely lost. The mutation gives some misfolding of protein that generally fails to produce fluoroscence, but sometimes also produce higher emission.

Green Fluoroscent Protein:

· The GFP was first discovered from the jellyfish Aequorea victoria. But it is also present in other organisms.

· The Aequorea victoria contains two luminescent proteins.

1) Aequorin

2) GFP, i.e. Green Fluoroscent Protein

· Aequorin, when interacts with Ca+2, it emits flashes of the blue light.

· The GFP aquires energy from Aequorin and emits green light.

Structure of GFP:

· The GFP from Aequorea victoria has an 11 stranded beta-barrel structure, with a alpha-helix running up the axis of the barrel.

· The chromophore is attached to alpha-helix in the center of barrel, few amino acids make the chromophore.

· The chromophore has Serine, Tyrosine and Glycine at the position 65; 66 and 67 respectively.

· 4-(p-hydroxybenzylidene)-imidazolidin-5-one attached to protein backbone through 1 and 2 position of the ring.

· Chromophore has hydrogen-bonding with amino acid residue and water molecules.