Electrospray ionisation mass spectrometry of biomolecular complexes

摘要: Electospray ionisation mass spectrometry (ESI-MS) has been used to study the noncovalent interactions in dsDNA-drug complexes, a protein-protein complex of DNA polymerase III E. coli and binding specificity cisplatin formation bifunctional adducts with DNA. ESI conditions have developed maximize detection dsDNA drugDNA complexes positive ion mode, comparable and/or superior results those achieved previously using negative ions. The gas phase stabilities different classes drugs examined. Different exhibit differences sequence selectivity, modify various ways are also known stabilise or destabilise dsDNA. ESI-MS/MS, desolvation temperature variation, in-source collision-induced dissociation experiments employed examine relative drug-dsDNA complexes. new class metallointercalators containing Ru examined by ESI-MS. Conditions were for optimal these mode their stoichiometries selectivity probed. order affinities was found correlate solution studies carried out related ruthenium compounds extended planar ligands dpq, dpqC dppz. ESI-MS supply “snapshot” components mixtures two proteins e186 θ, that form III. stability θ-e186 studied under instrumental conditions. behaviour this consistent significant contribution hydrophobic its stability, contrasted reported Tus-Ter complex. In addition, suggested θ subunit is involved stabilization e186. This earlier which shown e against thermal inactivation. Finally thesis at three well-characterized sites on ssDNA. A novel method better reaction purification ssDNA-cisplatin HPLC developed. 5′-GG-3′, 5′-AG-3′ 5′-GA-3′ characterized enzymatic digestion followed identification sites. ABBREVIATIONS (dpq) dipyrido[3,2-d:2′,3′-f]quinoxaline (dpqC) dipyrido[3,2-a:2′,3′-c](6,7,8,9-tetrahydro)phenazine (dppz) dipyrido[3,2-a:2′,3′-c]phenazine (phen) 1,10-phenanthroline adenine amu atomic unit bp base pairs C cytosine CD circular dichroism CID collision induced Da daltons DAPI 4′-6-diamidine-2-phenyl indole Dn daunomycin deoxyribonucleic acid double-stranded Dst distamycin Escherichia Electrospray Ionisation G guanine Hc Hoechst 33258 HMG high mobility group performance liquid chromatography m/z Mass-to-charge MALDI-MS matrix assisted laser desorption Mr molecular MS MS/MS tandem NMR nuclear magnetic resonance PA proton affinity Pd I phosphodiesterase II pol RNA ribonucleic ssDNA single-stranded T thymine Tm melting (of DNA)