Abstract: The tRNA anticodon loops always comprise seven nucleotides and is involved in many recognition processes with proteins and RNA fragments. We have investigated the nature and the possible interactions between the first (32) and last (38) residues of the loop on the basis of the available sequences and crystal structures. The data demonstrate the conservation of a bifurcated hydrogen bond interaction between residues 32 and 38, located at the stem/loop junction. This interaction leads to the formation of a non-canonical base-pair which is preserved in the known crystal structures of tRNA/synthetase complexes. Among the tRNA and tDNA sequences, 93 % of the 32.38 oppositions can be assigned to two families of isosteric base-pairs, one with a large (86 %) and the other with a much smaller (7 %) population. The remainder (7 %) of the oppositions have been assigned to a third family due to the lack of evidence for assigning them into the first two sets. In all families, the Y32.R38 base-pairs are not isosteric upon reversal (like the sheared G.A or wobble G.U pairs), explaining the strong conservation of a pyrimidine at position 32. Thus, the 32.38 interaction extends the sequence signature of the anticodon loop beyond the conserved U-turn at position 33 and the usually modified purine at position 37. A comparison with other loops containing both a singly hydrogen-bonded base-pair and a U-turn suggests that the 32.38 pair could be involved in the formation of a base triple with a residue in a ribosomal RNA component. It is also observed that two crystal structures of ribozymes (hammerhead and leadzyme) present similar base-pairs at the cleavage site.CI - Copyright 1999 Academic Press.