The early development and application of FTIR difference spectroscopy to membrane proteins: A personal perspective

Membrane proteins facilitate some of the most important cellular processes including energy conversion, ion transport and signal transduction. While conventional infrared absorption provides information about membrane protein secondary structure, a major challenge is to develop a dynamic picture of the functioning of membrane proteins at the molecular level. The introduction of FTIR difference spectroscopy around 1980 to study structural changes in membrane proteins along with a number of associated techniques including protein isotope labeling, site-directed mutagenesis, polarization dichroism, attenuated total reflection and time-resolved spectroscopy have led to significant progress towards this goal. It is now possible to routinely detect conformational changes of individual amino acid residues, backbone peptides, binding ligands, chromophores and even internal water molecules under physiological conditions with time-resolution down to nanoseconds. The advent of ultrafast pulsed-IR lasers has pushed this time-resolution down to femtoseconds. The early development of FTIR difference spectroscopy as applied to membrane proteins with special focus on bacteriorhodopsin is reviewed from a personal perspective.