chapter  12
10 Pages

Structure and Properties of GFP

In 1962, Osamu Shimomura and Frank Johnson published a paper describing the purification of


, a calcium-sensitive, chemiluminescent protein responsible for the bioluminescent properties of the jellyfish

Aequoria victoria

. When excited, this jellyfish emits green light at the fringes of its tentacles. Pure aequorin gives off a blue bioluminescence (emission peak 470 nm), but Shimomura and Johnson also found a companion protein “exhibiting a very bright, greenish fluorescence,” now called by the descriptive, if not very original, name of

green fluorescent protein

(GFP). This protein had an excitation peak close to 470 nm, suggesting that that the blue emission given off by aequorin is transferred to GFP via resonant energy transfer (Chapter 15) and GFP, in turn, emits the green glow characteristic of


. A fluorescent protein (FP) was an unusual concept at that time, which naturally

invited further investigation. It was found that GFP was encoded by a single gene, which was in due course sequenced. This was the beginning of a revolution in cell biology. GFP proved to be a very stable protein; its fluorescence is unaffected by harsh conditions such as 8 M urea, 1 percent SDS, and 2 days’ treatments with various proteases; it also survives aldehyde fixation. GFP is stable over a large range of pH (5.5 to 12) and at temperatures up to 65


C. Because it is so stable, is encoded by a single gene, needs no posttranscriptional modification, and requires no cofactors, GFP has revolutionized cell biology by providing an expressible marker that is directly visible in the fluorescence microscope.