When Ronald T. Raines at the University of Wisconsin, Madison, wanted to track peptides that traveled from the outside into mammalian cells, the biochemist found himself at a loss. He and coworkers needed a new tool--a molecule they could attach to the peptide that would fluoresce only inside the cell. They found the tool in a reaction used in drug delivery: the trimethyl lock.

Drug delivery strategists know that the crowding of three methyl groups in an o-hydroxycinnamic acid derivative causes strain that is relieved by cyclization--or locking--into a lactone. If a leaving group, such as an acetyl, is attached to the phenolic oxygen and a drug is placed at the carboxyl end through an amide bond, for example, cleavage of the leaving group (usually by an enzyme) leads to rapid cyclization and drug release.

In place of a drug, Raines and coworkers Sunil S. Chandran and Kimberly A. Dickson use fluorescent rhodamine 110. When rhodamine 110 is so bound, it doesn't fluoresce; when the trimethyl lock releases rhodamine 110, it fluoresces brightly [J. Am. Chem. Soc., published online Jan. 20, http://dx.doi.org/10.1021/ja043736v].

The new molecule is an especially good biological imaging tool, Raines says, because it is stable and not fluorescent in biological media until triggered. Inside the cell, esterases cleave the O-linked acetyl group and free the bright fluorophore.

The molecule can be conjugated to almost any protein. It can also be tailored to respond to "an enzyme of the user's choice," Raines says. Versions that fluoresce when a molecule moves into different cellular compartments--mitochondria, the Golgi apparatus, or the nucleus--are conceivable, he adds.

STRAINED When an esterase cleaves the O-R bond, the phenoxide, made more nucleophilic by strain from methyl crowding, attacks the amide carbon and frees fluorescent rhodamine 110.