Updating the sequence based classification of glycosyl hydrolases
The GH group includes several intronless GH genes of families 5, 6, 9, 10, 11, 16, and 26, three acetylxylan esterases ( interrupted by a large intron and with the canonical features of polyadenylation signal and intron-splicing boundaries of genes isolated from aerobic filamentous fungi (Durand et al.1995 ); a beta-succinyl-Co A synthetase ( gene, which clusters separately from other GH rumen fungi in figure 1 , can be explained by the highly biased amino acid content of the derived protein.On this basis, prokaryotic origins have been suggested for (Liu et al. However, these suggestions need a demonstration, and so, with this purpose in mind, we analyzed the G C content, the codon usage, and the phylogenetic trees derived from the multialignment of orthologous sequences worked out so far for rumen fungi genes.For analytical comparisons, we used gene sequences from strains that were homologous to some of the sequences from strain S85.Lignocellulosic material is hard and difficult to digest.Terrestrial herbivores might be expected to have a set of enzymes that can digest the cellulose and other polysaccharide structures of plants. Terrestrial animals have elaborated a different evolutive solution: symbiotic relationships with bacteria, protists, and fungi which carry out these activities in their own interests.As a measure of the distance between the codon usage of a gene (), we calculated the Hamming distance following a procedure similar to the one used for predicting protein structural classes (Chou and Zhang 1995 ).
To investigate the possible horizontal transfer of GH from rumen fungi, we aligned the catalytic domains of 54 endoglucanases of GH family 5 to a total length of 475 aa.
The strong similarity between different glycosyl hydrolases of rumen fungi and bacteria suggests that most, if not all, of the glycosyl hydrolases of rumen fungi that play an important role in the degradation of cellulose and other plant polysaccharides were acquired by horizontal gene transfer events.
This acquisition allows fungi to establish a habitat within a new environmental niche: the rumen of the herbivorous mammals for which cellulose and plant hemicellulose constitute the main raw nutritive substrate.
Because the codon usage patterns are species-specific, each gene in a genome tends to conform to the codon usage of its own species.
This allows us to define the null hypothesis that GH-encoding genes of rumen fungi should cluster with the remaining fungal chromosomal genes and not form a distinct group.