Expression, Purification, and Structure Determination of Human PTCH1-HH-N Complexes.

Patched-1 (PTCH1), a tumor suppressor, serves as the receptor of Hedgehog (HH) ligand and negatively regulates the HH signaling pathway. Mutations of PTCH1 are implicated in many human cancers. Structural investigation revealed the mechanism of PTCH1-mediated HH signal regulation, further facilitating the therapeutic development of cancers. Here, we describe the expression and purification of a nearly full-length functional PTCH1 variant, PTCH1*. With purified PTCH1* protein, two forms of PTCH1*-Sonic Hedgehog (SHH) complexes were assembled, and their structures subsequently determined by cryo-electron microscope (cryo-EM).