Nucleotide G-1207 of 18S rRNA is an essential component of the human 80S ribosomal decoding center.

mRNA analogues-derivatives of oligoribonucleotides consisting of two different codons and bearing an aryl azide group at the 5'-phosphates-were crosslinked to human 80S ribosomes by UV-irradiation of the various model complexes obtained in the presence of the cognate tRNAs. Three sequences, namely pUUUGUU (coding for Phe and Val), pUUCUAAA (first triplet coding for Phe and second being stop-codon), and pGUGUUU (coding for Val and Phe), have been used. Sequences of 18S rRNA containing nucleotides crosslinked to the mRNA analogues were examined by hydrolysis with RNase H in the presence of various cDNA probes.

Protein environment of mRNA at the decoding site of 80S ribosomes from human placenta as revealed from affinity labeling with mRNA analogs--derivatives of oligoribonucleotides.

Affinity labeling of 80S ribosomes from human placenta has been studied using various mRNA analogs, namely, 2',3'-O-[4-(N-2-chloroethyl)-N-methylamino]benzylidene derivatives of oligoribonucleotides (Up)(n-1)U[32P]pC (n = 3, 6 or 12) and AUGU3[32P]pC as well as ([4-(N-2-chloroethyl)-N-methylamino]benzylmethyl-[5'-32P]-phospham ide derivatives of pAUGUn (n = 3 or 6). Labeling of 80S ribosomes with the derivatives of oligouridylates was carried out in complexes obtained nonenzymatically in the presence of saturating amounts of Phe-tRNA(Phe). Complexes with derivatives bearing AUG codon were obtained using a fractionated lysate from rabbit reticulocytes which contained protein translation factors and was deprived from endogeneous ribosomes and mRNAs.