Lens crystallins were isolated from the homogenates of mammalian eye lenses derived from three different species by gel permeation chromatography and characterized by SDS-gel electrophoresis, isoelectric focusing, amino acid analysis and N-terminal sequence analysis. Five fractions corresponding to HMα-,α-,βH-,βL-andγ-crystallins were obtained for the crystallins from these phylogenetically distant species. The native molecular masses for these purified fractions and their polypeptide compositions were determined by gel filtration and SDS-gel electrophoresis respectively, revealing the typical subunit compositions for each classified crystallin. The gel pattern ofγ-crystallins from the marmot lens appeared to be more complex than those of gibbon and deer lenses. Comparison of the amino acid contents of each orthologous class of mammalian crystallins with those of evolutionarily distant species still exhibited similarity in their amino acid compositions. The charge heterogeneity of each crystallin fraction can be detected by isoelectric focusing under denaturing conditions. N-terminal sequence analysis of the crystallin fractions revealed that all fractions except that ofγ-crystallin are N-terminally blocked. Extensive sequence similarity between mammalianγ-crystallin polypeptides were found, which suggested the close relatedness ofγ-crystallins amongst different species of mammals and also established the heterogeneous nature of this multigene family.