Chitons are ancient polyplacophora mollusk covered with biominerals that may shed light on the origin and evolution of biomineralization. While the body plan of chitons is strictly conservative since the Cambrian, the conservation degree of biomineralized mechanisms among chitons are unclear. Hence, we extracted the matrix proteins of two biominerals (shell plates and spicules) from Acanthochitona rubrolineata, which is widely distributed in the coastal region of China. Liquid chromatography coupled to tandem mass spectrometry was utilized to obtain their proteome information, and proteomic analysis was performed with the species-specific transcriptome library. By comparing biomineralization proteins of Acanthopleura loochooana from the southeastern coast of China, the von Willebrand factor type A domain, chitin-binding domain, ferritin, and cadherin are found to be shared in both species. This study provides an important reference for understanding the evolutionary conservatism on chiton biomineralization.