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Zoonomia is one of the largest comparative genomics resources produced to date. By comparing the genomes of 240 mammal species, we harness over 100 million years of evolution to explore genome function. Overall at least 11% of the human genome is constrained, and constrained bases are enriched for variants explaining common disease heritability more than any other functional annotation. We associate coding and regulatory variants with variation in hibernation and brain size across placental mammals. Nearly half of highly constrained bases are unannotated in existing data resources, highlighting how much of the regulatory landscape of the genome has yet to be explored.