Novel amidinating cross-linker for facilitating analyses of protein structures and interactions.
A novel bifunctional thioimidate cross-linking reagent (diethyl suberthioimidate) that modifies amines without sacrificing their native basicity is developed. Intermolecular cross-linking of neurotensin and intramolecular cross-linking of cytochrome c under physiological conditions is investigated with this reagent. Because it does not perturb the electrostatic properties of a protein, it is unlikely to lead to artifactual conclusions about native protein structure. The interpeptide cross-links formed with this reagent are easily separated from other tryptic fragments using strong cation exchange chromatography, and they have a readily identified mass spectrometric signature. The use of this novel amidinating protein cross-linking reagent holds great promise for efficient, large-scale structural analysis of complex systems.
Lauber
Matthew A
MA
Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA.
Reilly
James P
JP
D000577:Amides / Q000737:chemistry*; D000588:Amines / Q000737:chemistry; D000818:Animals; D002417:Cattle; D002852:Chromatography, Ion Exchange; D003432:Cross-Linking Reagents / Q000138:chemical synthesis* / Q000737:chemistry; D045304:Cytochromes c / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007096:Imidoesters / Q000138:chemical synthesis* / Q000737:chemistry; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000737:chemistry
D000577:Amides; D000588:Amines; D003432:Cross-Linking Reagents; D007096:Imidoesters; D011506:Proteins; D013438:Sulfhydryl Compounds; D045304:Cytochromes c