The observation that homology is often manifested as sequence similarity suggests that the evolutionary pathway relating the members of a family of proteins may be deduced by examination of sequence similarity. This approach is based on the notion that sequences that are more similar to one another have had less evolutionary time to diverge than have sequences that are less similar. This method can be illustrated by using the three globin sequences in Figures 6.11 and 6.13, as well as the sequence for the human hemoglobin β chain. These sequences can be aligned with the additional constraint that gaps, if present, should be at the same positions in all of the proteins. These aligned sequences can be used to construct an evolutionary tree in which the length of the branch connecting each pair of proteins is proportional to the number of amino acid differences between the sequences (Figure 6.21).

Such comparisons reveal only the relative divergence times—for example, that myoglobin diverged from hemoglobin twice as long ago as the α chain diverged from the β chain. How can we estimate the approximate dates of gene duplications and other evolutionary events? Evolutionary trees can be calibrated by comparing the deduced branch points with divergence times determined from the fossil record. For example, the duplication leading to the two chains of hemoglobin appears to have occurred 350 million years ago. This estimate is supported by the observation that jawless fish such as the lamprey, which diverged from bony fish approximately 400 million years ago, contain hemoglobin built from a single type of subunit (Figure 6.22). These methods can be applied to both relatively modern and very ancient molecules, such as the ribosomal RNAs that are found in all organisms. Indeed, such an RNA sequence analysis led to the realization that Archaea are a distinct group of organisms that diverged from Bacteria very early in evolutionary history.

184

Evolutionary trees that encompass orthologs of a particular protein across a range of species can lead to unexpected findings. For example, let us consider the unicellular red alga Galdieria sulphuraria, a remarkable eukaryote that can thrive in extreme environments, including at temperatures up to 56°C, at pH values between 0 and 4, and in the presence of high concentrations of toxic metals. G. sulphuraria belongs to the order Cyanidiales, clearly within the eukaryotic branch of the evolutionary tree (Figure 6.23A). However, the complete genome sequence of this organism revealed that nearly 5% of the G. sulphuraria ORFs encode proteins that are more closely related to bacterial or archaeal, not eukaryotic, orthologs. Furthermore, the proteins that exhibited these unexpected evolutionary relationships possess functions that are likely to confer a survival advantage in extreme environments, such as the removal of metal ions from inside the cell (Figure 6.23B). One likely explanation for these observations is horizontal gene transfer, or the exchange of DNA between species that provides a selective advantage to the recipient. Amongst prokaryotes, horizontal gene transfer is a well-characterized and important evolutionary mechanism. For example, as we shall discuss in Chapter 9, exchange of plasmid DNA between bacterial species likely facilitated the acquisition of restriction endonuclease activities. However, recent studies such as those on G. sulphuraria, made possible by the expansive growth of complete genome sequence information, suggest that horizontal gene transfer from prokaryotes to eukaryotes, between different domains of life, may also represent evolutionarily significant events.

185