Identification of Chimeric αβγ Diterpene Synthases Possessing both Type II Terpene Cyclase and Prenyltransferase Activities.

Abstract

Two unusual diterpene synthases composed of three domains (α, β, and γ) were identified from fungal Penicillium species. They are the first enzymes found to possess both type II terpene cyclase (TC) and prenyltransferase (PT) activities. These enzymes were characterized by heterologous expression in Aspergillus oryzae and in vitro experiments with wild-type, mutated, and truncated enzymes. The results revealed that the α domain in the C-terminal region of these enzymes was responsible for the PT activity, whereas the βγ domains in the N-terminal region composed the type II TC, and formed copalyl diphosphate (2). Additionally, between the α and βγ domains, there is a characteristic linker region, in which minimal secondary structure is predicted. This linker does not exist in the characterized three-domain (αβγ) terpene synthases known as monofunctional type I or type II TCs, or bifunctional type I and type II TC enzymes. Therefore, both the catalytic activities and protein architecture substantially differentiate these new enzymes from the previously characterized terpene synthases.