Figure 8. Clustering of ENC values for Hsp60 genes from Chordata. Cluster #1 includes the ENC values of Hsp60 genes of Mammalia, Aves, Reptilia, and Amphibia. Cluster #2 includes ENC values of Hsp60 genes of Fish. Clustering was performed using the value of the GC3 content corresponding to the equilibrium pointEp , which was determined earlier (see GC-content and mutation pressure for codon usage).
As can be seen, the ENC values of the Hsp60 genes from Chordata form two groups (Figure 8). Hsp60 genes from cluster #1 (GC3 is less than Ep ) belong to Mammalia, Aves, Reptilia, and Amphibia. In turn, cluster #2 (GC3 is larger thanEp ) consists only of Hsp60 genes from Actinopterygii (ray-finned fishes). The ENC values and the GC3 content of the Hsp60 genes in cluster #2 differ more than in cluster #1. This feature can be explained by the fact that Hsp60 from Fish is evolutionarily far from Hsp60 from other Chordata classes14.
It should be noted that the ENC value of the gene can correlate with the level of its expression in the cell59–62. Based on current research, the following ENC thresholds have been established for determining the level of expression of the Hsp60 gene: ENC < 40 for highly expressed genes59,63,64; 40 < ENC ≤ 55 for moderately expressed genes59,63,65; ENC > 55 for lowly expressed genes60,63. These conditions can be used to assess the main trends in the average codon usage bias for Hsp60 genes (Figure 9).