Joel S. Hayflick

Mutagenesis of the three bases preceding the start codon of the beta-galactosidase mRNA and its effect on translation in Escherichia coli.

The effect on the translation efficiency of various mutations in the three bases (the ‐1 triplet) that precede the AUG start codon of the beta‐galactosidase mRNA in Escherichia coli was studied. Of the 39 mutants examined, the level of expression varies over a 20‐fold range. The most favorable combinations of bases in the ‐1 triplet are UAU and CUU. The expression levels in the mutants with UUC, UCA or AGG as the ‐1 triplet are 20‐fold lower than those with UAU or CUU. In general, a U residue immediately preceding the start codon is more favorable for expression than any other base; furthermore, an A residue at the ‐2 position enhances the translation efficiency in most instances. In both cases, however, the degree of enhancement depends on its context, i.e. the neighboring bases. Although the rules derived from this study are complex, the results show that mutations in any of the three bases preceding the start codon can strongly affect the translational efficiency of the beta‐galactosidase mRNA.

The amino acid sequence of murine p53 determined from a c-DNA clone

A c-DNA clone containing the complete sequence information for the murine p53 protein, from embryonal carcinoma cells, has been isolated. The nucleotide sequence of this clone reveals an open reading frame encoding a protein of 390 amino acids with a molecular weight of 43,364 Da. The NH2-terminal domain of this protein is acidic whereas the carboxyl terminus is rich in basic amino acid residues. These terminal domains are separated by a proline-rich, hydrophobic run of amino acids. Proline comprises approximately 10% of the total amino acid residues. Two tryptic peptides, derived from p53 protein radiolabeled with either methionine or proline, were purified and the position of these labeled residues in the peptide was determined. The positions of three methionine and five proline residues in these two peptides matched the amino acid sequence of the predicted open reading frame determined from the c-DNA clone.