The unusual diterpene (C20) synthase copalyl diphosphate synthase from Penicillium verruculosum (PvCPS) is the first bifunctional terpene synthase identified with both prenyltransferase and class II cyclase activities in a single polypeptide chain with αβγ domain architecture. The C-terminal prenyltransferase αdomain generates geranylgeranyl diphosphate which is then cyclized to form copalyl diphosphate at the N-terminal βγ domain interface. We now demonstrate that PvCPS exists as a hexamer at high concentrations - a unique quaternary structure for known αβγ terpene synthases. Hexamer assembly is corroborated by a 2.41 Å-resolution crystal structure of the α domain prenyltransferase obtained from lim... More
The unusual diterpene (C20) synthase copalyl diphosphate synthase from Penicillium verruculosum (PvCPS) is the first bifunctional terpene synthase identified with both prenyltransferase and class II cyclase activities in a single polypeptide chain with αβγ domain architecture. The C-terminal prenyltransferase αdomain generates geranylgeranyl diphosphate which is then cyclized to form copalyl diphosphate at the N-terminal βγ domain interface. We now demonstrate that PvCPS exists as a hexamer at high concentrations - a unique quaternary structure for known αβγ terpene synthases. Hexamer assembly is corroborated by a 2.41 Å-resolution crystal structure of the α domain prenyltransferase obtained from limited proteolysis of full-length PvCPS, as well as the ab initio modeling of full-length PvCPS derived from small-angle X-ray scattering data. Hexamerization of the prenyltransferase αdomain appears to drive the hexamerization of full-length PvCPS. The PvCPS hexamer dissociates into lower-order species at lower concentrations, as evidenced by size-exclusion chromatography in-line with multiangle light scattering, sedimentation velocity analytical ultracentrifugation, and native polyacrylamide gel electrophoresis experiments, suggesting that oligomerization is concentration dependent. Even so, PvCPS oligomer assembly does not affect prenyltransferase activity in vitro.,Copyright © 2020 Elsevier Inc. All rights reserved.
