Robert C. Pini
A light-activated reagent that can bind to protein molecules and then sever them when irradiated by a 344-nm light could offer microbiologists a versatile tool for exploring the structure and behavior of proteins. Scientists could use this tool to sequence and manipulate proteins, controlling the reagent's scissorlike effect precisely, in part because the reagent can be selectively activated and in part because the molecule will bind only in specific sites on the protein.
The reagent was designed and studied by researchers at the University of Connecticut in collaboration with Glaxo Wellcome in Research Triangle, N.C., and Columbia University in New York. "So far we looked at several wavelengths from 310 to 380 nm, and the cleavage efficiency varies. The molelcule absorbs light very strongly at [the 344 nm] wavelength," said Challa V. Kumar, an associate professor of chemistry at the University of Connecticut.
The designer molecule, N-(phen-ylalanine)-4-(1-pyrene)butyramide, or Py-Phe, binds in specific sites. The pyrene part is hydrophobic, and the phenylalanine carboxyl part is hydrophilic, so Py-Phe will bind on a protein only where a hydrophobic site is adjacent to a hydrophilic one.
Chemists and biologists speculate that the molecule can be used as a general reagent for cutting proteins, which would make it useful for molecular studies of proteins and peptides. Researchers have shown that a Py-Phe molecule can bind to lysozyme and albumin proteins and split them cleanly in two.
"Many techniques can cut a protein with enzymes thermally," said Miguel Garcia Garibay, associate professor of chemistry at UCLA. But such methods make no distinction between events, so are suitable only for studying material in steady state.
"This technique can be activated precisely, so it has applications in genetics to study processes of [molecular] interaction within a defined time span," Garibay said.
Kumar expects to begin tests on streptavidin, cholesterol oxidase and other key proteins involved in human disease states. "If we can selectively photodestruct key proteins, then we may target cancer cells or specific pathogens," he said.
To activate the protein scissors, researchers used a lamp and a monochromator. Light raises the Py-Phe radical to an excited state, initiating a sequence of molecular reactions that split the peptide bond.