An information analysis of the 5' (donor) and 3' (acceptor) sequences spanning the ends of nearly 1800 human introns has provided evidence for structural features of splice sites that bear upon spliceosome evolution and function: (1) 82% of the sequence information (i.e. sequence conservation) at donor junctions and 97% of the sequence information at acceptor junctions is confined to the introns, allowing codon choices throughout exons to be largely unrestricted. The distribution of information at intron-exon junctions is also described in detail and compared with footprints. (2) Acceptor sites are found to possess enough information to be located in the transcribed portion of the human genome, whereas donor sites possess about one bit less than the information needed to locate them independently. This difference suggests that acceptor sites are located first in humans and, having been located, reduce by a factor of two the number of alternative sites available as donors. Direct experimental evidence exists to support this conclusion. (3) The sequences of donor and acceptor splice sites exhibit a striking similarity. This suggests that the two junctions derive from a common ancestor and that during evolution the information of both sites shifted onto the intron. If so, the protein and RNA components that are found in contemporary spliceosomes, and which are responsible for recognizing donor and acceptor sequences, should also be related. This conclusion is supported by the common structures found in different parts of the spliceosome.