Anion binding by fluorescent Fmoc-protected amino acids

Two non-natural amino acids with fluorescent urea side-chains were prepared from Fmoc-protected aspartic and glutamic acids. In acetonitrile solution, the emission of the Asp derivative is strongly quenched by HCO3- or H2PO4- (K10(4) M-1) but not by less-basic Cl- or NO3-. Solutions containing excess bicarbonate ion appear peach-colored, with (abs) at 394 and 495nm ascribed to the anion complex and urea-deprotonated sensor, respectively. Corresponding fluorescence bands are observed at 475 and 579nm. Dihydrogenphosphate is not sufficiently basic to remove H+ from the ground state of the fluorophore. However, deprotonation of the excited state occurs in the presence of>1 equiv of H2PO4- ((em)=578nm). According to H-1 NMR in DMSO-d(6), recognition of H2PO4- occurs at the urea N-H groups and the amino acid backbone N-H. DFT techniques further predict that the backbone C=O group accepts an H-bond from the anion. The Glu derivative has lower affinity for anions; the additional CH2 group in its side-chain apparently sets the backbone N-H and C=O too far from the urea to contribute significantly to binding. To demonstrate suitability for standard Fmoc-based solid-phase peptide synthesis, the Asp derivative was incorporated into a 12-residue peptide.