title: A novel method for detection of selenomethionine incorporation in protein crystals via Raman microscopy.
abstract: Multiwavelength anomalous dispersion (MAD) is the most widespread approach in structural biology for the determination of the crystal structure of a novel protein. Mass spectrometry is currently used to evaluate the selenomethionine (SeMet) content in solution, but excluding fluorescence spectroscopy at the absorption edge, no other routine method to check the SeMet incorporation and storage in the crystal state is yet available. Raman microscopy is an increasingly popular tool in physical biochemistry, with applications ranging from studies of ligand binding to secondary-structure analysis. Here, a novel methodological development is presented for the analysis via Raman microscopy of SeMet-labelled protein crystals to be used for MAD crystallography. The method is described and supported by validation and application to two novel proteins (a betagamma-crystallin-like protein and a DNA-binding protein). Markers of the SeMet residues are in the range 570-600 cm(-1), where proteins do not usually show Raman bands.
authors.affiliation: Dipartimento di Chimica, Università degli Studi di Napoli 'Federico II', Complesso Monte S. Angelo, Via Cinthia, I-80126 Napoli, Italy.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Vergara
authors.forename: Antonello
authors.initials: A
authors.lastname: Merlino
authors.forename: Elio
authors.initials: E
authors.lastname: Pizzo
authors.forename: Giuseppe
authors.initials: G
authors.lastname: D'Alessio
authors.forename: Lelio
authors.initials: L
authors.lastname: Mazzarella
mesh_terms: D001572:Benzofurans / Q000737:chemistry; D018360:Crystallography, X-Ray / Q000379:methods; D011506:Proteins / Q000737:chemistry*; D012645:Selenomethionine / Q000737:chemistry*; D013059:Spectrum Analysis, Raman / Q000379:methods*
chemical_list: D001572:Benzofurans; D011506:Proteins; C052047:geodin; D012645:Selenomethionine