S63 Use of NMR spectroscopy and nanospray mass spectrometry to characterise binding of lead compounds for drug design in  1-antitrypsin deficiency

Pathogenic mutations in the gene for α1 -antitrypsin predispose to aberrant conformational transitions of the translated protein molecules resulting in their self-association to form polymer chains. Polymerisation causes circulating deficiency of α1 -antitrypsin while predisposing to hepatic cirrhosis and severe, early-onset emphysema. Targeting the conformational transitions underlying polymerisation via ligand binding and stabilisation of the physiological native state is therefore a goal of drug design in α1 -antitrypsin deficiency. To complement previous structure-led approaches we have developed NMR spectroscopy and nanospray mass spectrometry as medium-throughput screening tools for such ligands. The coupling of these techniques combines highly sensitive detection of ligand binding with assessment of binding sites, stoichiometry, cooperativity and binding constants. We have used the TTAI peptide, developed within an existing programme of drug design, as a test case. The data demonstrate highly co-operative, slow, tight binding of two copies of the peptide in adjacent parts of the α1 -antitrypsin molecule. TTAI peptide binding is shown to induce widespread conformational change all over the molecule with the exception of β-sheet C. These data prove the utility of NMR spectroscopy and nanospray mass spectrometry in characterising ligand binding whilst providing a highly detailed template for use in specific screening for TTAI peptide-mimetic compounds.