Identification of a novel amino acid, o-bromo-L-phenylalanine, in egg-associated peptides that activate spermatozoa

Eight sperm-activating peptides containing a novel amino acid were isolated from the egg jelly of the sea urchin Tripneustes gratilla. Accurate mass measurement of the peptide in FAB mass spectrometry showed that the mass of the novel amino acid residue was 224.978. On the basis of the isotopic ion distribution and the degree of unsaturation, the mass value indicated that the elemental composition of the amino acid residue was C9H8O1N1Br1, suggesting that the novel amino acid was bromophenylalanine. Proton NMR spectroscopy, amino acid analysis, and RP-HPLC with three synthetic isomers of bromophenylalanine demonstrated that o-bromophenylalanine was the novel amino acid. Derivatization of the amino acid with Marfey's reagent, (1-fluoro-2,4-dinitrophen-5-yl)-L-alanine amide (FDAA), further indicated that the amino acid was the L-isomer. In other sperm-activating peptides isolated from the egg jelly of the sea urchin, both m- and p-bromophenylalanines were discovered. The presence of m-bromophenylalanine has not been previously reported in natural products, while p-bromophenylalanine is found in theonellamide F, an antifungal bicyclic peptide from a marine sponge.