Project description:Gigantopithecus blacki was a giant hominid that inhabited Southeast Asia during the Pleistocene. Its evolutionary relationship to other great ape species, and their divergence during the Middle and Late Miocene (16-5.3 Mya), remains disputed. In part, this is due to the absence of cranial and postcranial remains and size-induced allometry. Proposed hypothesis on the phylogenetic positions of Gigantopithecus have therefore been wide-ranging among hominoids, but none has received independent validation based on molecular evidence. To clarify the phylogenetic placement of Gigantopithecus blacki, we retrieved enamel proteome sequences from a 1.9 million years (Mya) old molar found in Chuifeng Cave, China. We demonstrate that Gigantopithecus is most closely related to orangutans (genus Pongo). We also estimate the Gigantopithecus-Pongo divergence to about 10-12 Mya, implying its speciation is part of the Miocene radiation of great apes. These sequences are approximately 6 times older, in a normalized thermal context, than any previously published mammalian proteome or genome. The survival of an Early Pleistocene dental enamel proteome in the subtropics further expands the scope of palaeoproteomic analysis into geographic areas and time periods previously considered incompatible with biomolecular preservation.