A 20-AMINO-ACID AUTONOMOUS RNA-BINDING DOMAIN CONTAINED IN AN ENOYL-COA HYDRATASE

摘要: A+U-rich elements in the 3' untranslated region of mRNA species coding for lymphokines and early response genes play a pivotal role control their rapid turnover. In search corresponding trans-acting factors, we have previously affinity-purified cloned human 32-kDa A+U-binding protein, termed AUH. AUH exhibited dual activities, namely A+U-specific RNA-binding catalytic activity as enoyl-CoA hydratase. this report map site by analysis series deletion substitution recombinant proteins. Ultraviolet cross-linking experiments demonstrated that 20-amino-acid segment, Lys109-Ile128, abolished more than 80% relative activity. This segment conferred when fused to maltose binding protein. Binding fusion protein RNA was significantly competed an AUUUA cluster poly(U), followed poly(G), but not poly(A) nor poly(C). Furthermore, synthetic peptide Lys109-Ile128. Circular dichroic measurement indicated formation specific complex between poly(U) with poly(A). The identified 20 amino acids therefore constitute automonous domain, distinct from RNA-recognition motifs family ribonucleoproteins or NAD/RNA-binding sites dehydrogenases found hitherto reported Replacement Arg125 motif Glu reduced twofold, indicating residue is integral function. Deletion other parts did impair any significant extent. By contrast, hydratase required intact three-dimensional conformation, most mutations downstream Ser68 impaired enzymatic