The gene of an archaeal a-l-fucosidase is expressed by translational frameshifting

Cobucci-Ponzano, Beatrice; Conte, Fiorella; Benelli, Dario; Londei, Paola; Flagiello, Angela; Monti, Maria; Pucci, Piero; Rossi, Mosè; Moracci, Marco

Remarkably, the MALDIMS analysis of the products of the wild-type fucA1 gene revealed the presence of a second Peptide B at m/z 1258.6 that is absent in the spectra of the Ssa-fuc control protein ( Figure 2B and C). The sequence of Peptide B obtained by LCMSMS ( Figure 2D ) was Lys-Phe-Gly-Pro-Val-Thr-Asp-Phe-Gly-Tyr-Lys. This sequence differs only by one amino acid from Peptide A demonstrating that the interrupted gene fucA1 expresses in E.coli two fulllength proteins originated by different À1 frameshifting events. Polypeptide A results from a shift in a site A and it is identical to Ssa-fuc prepared by site-directed mutagenesis (24) , suggesting that the expression occurred with the simultaneous P-and A-site slippage. Instead, polypeptide B, named Ssa-fuc B , is generated by frameshifting in a second site B as the result of a single P-site slippage ( Figure 2E) .