The pentaammineruthenium(III)histidine complex in ribonuclease A: Application to the assignment of histidine proton resonances

✍ Scribed by C.Robert Matthews; Joanne Recchia; Claudia L. Froebe

• Publisher: Elsevier Science
• Year: 1981
• Tongue: English
• Weight: 766 KB
• Volume: 112
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(81)90301-8

✦ Synopsis

The assignment of two histidine proton resonances in the proton NMR spectrum of ribonuclease A has been made by forming a paramagnetic complex between pentaammineruthenium(II1) and the N-3 nitrogen of a single histidine residue. Reaction of chloropentaammineruthenium(III)dichloride with ribonuclease A in 0.1 M Tris-HCI, pH 7.0, 25°C yields a variety of products in which various histidine residues have been labeled. Cation-exchange chromatography affords the isolation of a specific derivative, labeled at a single histidine residue, that retains 66% of the activity toward the hydrolysis of 2',3'-cyclic CMP. The site of labeling was determined by peptide mapping to be histidine 105. The binding of ruthenium results in the disappearance of both a histidine C-2 and a C-4 proton resonance from the downfield region of the proton NMR spectrum, as expected from model compound studies. The assignment of these two resonances to histidine 105 is in agreement with a previous assignment (J. L. Markley, 1975, Biochemistry 14, 3546-3554), thereby demonstrating the potential utility of this ruthenium reagent in the assignment of histidine resonances in the proton NMR spectra of other proteins.