SIDE-CHAIN SELECTIVE AND COVALENT LABELING OF PROTEINS BY ORGANOMETALLIC COMPLEXES OF HEAVY TRANSITION-METALS - POSSIBLE APPLICATION IN RADIO-CRYSTALLOGRAPHY OF PROTEINS

Organo-rhenium and organo-tungsten N-succinimidyl and N-sulfosuccinimi dyl esters react at amino groups of proteins to form stable amide bond s between the protein and the heavy transition metal complexes. We sho w here that factors such as pH of the reaction medium and nature of th e reagent (coordinating metal, surrounding ligands, type of ester) had a marked influence on the final number of coupled organometallic grou ps per protein molecule, defined as the coupling ratio. By operating w ith stoichiometric quantities of reagent with respect to the number of amino groups, up to 30 organometallic groups were introduced into the model protein, bovine serum albumin (BSA). BSA conjugates were charac terized by gel electrophoresis in non-denaturating conditions. Hen egg -white lysozyme organo-tungsten conjugates were prepared in the same m anner and the peptides resulting from their tryptic hydrolysis were an alyzed by reverse-phase HPLC. A tentative assignment of the preferenti al amino sites of conjugation is suggested from the latter results.