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Figure 9.12. Sequence comparison of galactokinases from different species. (a) The galactokinase reaction. (b) Comparison of a region of human galactokinase (amino acids 35-54 of the 392 amino acid protein). Key: Hs - Homo sapiens, Sc - Saccharomyces cerevisiae, Ec - Escherichia coli, Bs - Bacillus subtilis, Ca - Candida albicans, Hi - Haemophilus influenzae, St - Salmonella typhimurium, Kl - Kluyveromyces lactis, At - Arabidopsis thaliana. Amino acids have been coloured according to their properties. Blue indicates positively charged amino acids (H, K, R), red indicates negatively charged residues (D, E), green indicates polar neutral residues (S, T, N, Q), grey indicates non-polar aliphatics (A, V, L, I, M) and purple indicates non-polar aromatic residues (F, Y, W). Brown is used to indicate proline and glycine, while yellow indicates cysteine homologous recombination is used in both yeast and in higher-eukaryotic cells to disrupt the functional copy of a gene within a genome. The phenotype of the disrupted mutant can then be assessed in order to attempt to identify the natural function of the wild-type gene. This approach works well for many genes. For example, the previously uncharacterized yeast gene SNU17 shows little similarity to other proteins when compared using database searches. A yeast strain knocked out for SNU17, however, shows a slow-growth phenotype and is defective in pre-mRNA splicing (Gottschalk et al., 2001), indicating that the protein is involved in the splicing process. The difficulty with this approach is that, often, the deleted strain is either non-viable or is indistinguishable from the wild-type. Neither of these outcomes makes functional assignment possible - the non-viable state suggests that the protein may be playing a vital role in the cell, but may not yield any further clues to that role. An alternative approach to gene assignment is to overproduce a protein, by carrying the gene on a high-copy-number plasmid, to attempt to observe a phenotype.

Despite the availability of the techniques described above, much of the assignment of gene function must be performed on an individual gene basis. This remains a large task in an experimentally tractable organism for the 2000 or so unidentified yeast genes, but the complete identification of the 30 000 or so human genes seems daunting.

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