RAPID PROTEIN SEQUENCING BY TANDEM MASS-SPECTROMETRY AND CDNA CLONINGOF P20-CGGBP - A NOVEL PROTEIN THAT BINDS TO THE UNSTABLE TRIPLET REPEAT 5'-D(CGG)(N)-3' IN THE HUMAN FMR1 GENE

The autonomous expansion of the unstable 5'-d(CGG)(n)-3' repeat in the 5'-untranslated region of the human FMR1 gene leads to the fragile X syndrome, one of the most frequent causes of mental retardation in hum an males, We have recently described the isolation of a protein p20-CG GBP that binds sequence-specifically to the double stranded trinucleot ide repeat 5'-d(CGG)-3' (Deissler, H., Behn-Krappa, A., and Doerfler, W. (1996) J. Biol. Chem. 271, 4327-4334), We demonstrate now that the p20-CGGBP can also bind to an interrupted repeat sequence, Peptide seq uence tags of p20-CGGBP obtained by nanoelectrospray mass spectrometry were screened against an expressed sequence tag data base, retrieving a clone that contained the full-length coding sequence for p20-CGGBP. A bacterially expressed fusion protein p20-CGGBP-6xHis exhibits a bin ding pattern to the double-stranded 5'-d(CGG),-3' repeat similar to th at of the authentic p20-CGGBP, This novel protein lacks any overall ho mology to other known proteins but carries a putative nuclear localiza tion signal, The p20-CGGBP gene is conserved among mammals but shows n o homology to non-vertebrate species, The gene encoding the sequence f or the new protein has been mapped to human chromosome 3.