title: Towards a rational approach for heavy-atom derivative screening in protein crystallography.
abstract: Heavy-atom derivatization is routinely used in protein structure determination and is thus of critical importance in structural biology. In order to replace the current trial-and-error heavy-atom derivative screening with a knowledge-based rational derivative-selection method, the reactivity of more than 40 heavy-atom compounds over a wide range of buffer and pH values was systematically examined using peptides which contained a single reactive amino-acid residue. Met-, Cys- and His-containing peptides were derivatized against Hg, Au and Pt compounds, while Tyr-, Glu-, Asp-, Asn- and Gln-containing peptides were assessed against Pb compounds. A total of 1668 reactive conditions were examined using mass spectrometry and were compiled into heavy-atom reactivity tables (http://sis.niaid.nih.gov/cgi-bin/heavyatom_reactivity.cgi). The results showed that heavy-atom derivatization reactions are highly linked to buffer and pH, with the most accommodating buffer being MES at pH 6. A group of 21 compounds were identified as most successful irrespective of ligand or buffer/pH conditions. To assess the applicability of the peptide heavy-atom reactivity to proteins, lysozyme crystals were derivatized with a list of peptide-reactive compounds that included both known and new compounds for lysozyme derivatization. The results showed highly consistent heavy-atom reactivities between the peptides and lysozyme.
authors.affiliation: Structural Immunology Section, Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 12441 Parklawn Drive, Rockville, Maryland 20852, USA.
authors.forename: Johnson
authors.initials: J
authors.lastname: Agniswamy
authors.forename: M Gordon
authors.initials: MG
authors.lastname: Joyce
authors.forename: Carl H
authors.initials: CH
authors.lastname: Hammer
authors.forename: Peter D
authors.initials: PD
authors.lastname: Sun
mesh_terms: D000596:Amino Acids / Q000737:chemistry; D003461:Crystallography / Q000379:methods*; D005583:Fourier Analysis; D007854:Lead / Q000737:chemistry; D008024:Ligands; D013058:Mass Spectrometry; D019216:Metals, Heavy / Q000737:chemistry*; D008958:Models, Molecular; D008968:Molecular Conformation; D009113:Muramidase / Q000737:chemistry; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry; D011506:Proteins / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: D000596:Amino Acids; D008024:Ligands; D019216:Metals, Heavy; D010455:Peptides; D011506:Proteins; D007854:Lead; D009113:Muramidase