Protein structure-function relationship studied by single crystal polarized absorption microspectrophotometry

Abstract

Are the structure and function of a protein in the crystal the same as in solution? The question, put forward at the first publications of the three dimensional structure of proteins [1], is still dramatically valid. In fact, about ten thousand protein structures have been determined by X-ray crystallography, and more are exponentially coming out, but only 5% of these proteins have been tested whether their functional properties are maintained in the crystal. There is no a priori answer and each protein has to be individually investigated. A powerful technique for the determination of functional properties of proteins in the crystal is polarized absorption microspectrophotometry [2]. The chromophoric properties of cofactors, coenzymes, ligands, substrates and substrate analogs are exploited to monitor function-related spectroscopic properties and to determine kinetic and thermodynamic parameters. Two case-studies, crystals of T state hemoglobin and pyridoxal 5′-phosphate-dependent O-acetylserine sulfhydrylase, will allow to evidence some of the advantages of carrying out functional studies in the same physical state where the three dimensional structure is obtained.