Crystallization and X-ray diffraction analysis of a putative bacterial class I labdane-related diterpene synthase
Labdane-related diterpenoids are natural products with potential pharmaceutical applications that are rarely found in bacteria. Here, a putative class I labdane-related diterpene synthase (LrdC) identified by genome mining in a streptomycete was successfully crystallized using the microbatch method. Crystals of the LrdC enzyme were obtained in a holo form with its natural cofactor Mg2+(LrdC-Mg2+) and in complex with inorganic pyrophosphate (PPi) (LrdC-Mg2+–PPi). Crystals of native LrdC-Mg2+diffracted to 2.50 Å resolution and belonged to the trigonal space groupP3221, with unit-cell parametersa = b = 107.1,c= 89.2 Å. Crystals of the LrdC-Mg2+–PPicomplex grown in the same conditions as the native enzyme with PEG 8000 diffracted to 2.36 Å resolution and also belonged to the trigonal space groupP3221. Crystals of the LrdC-Mg2+–PPicomplex grown in a second crystallization condition with PEG 3350 diffracted to 2.57 Å resolution and belonged to the monoclinic space groupP21, with unit-cell parametersa= 49.9,b= 104.1,c= 66.5 Å, β = 111.4°. The structure was determined by the single-wavelength anomalous dispersion (SAD) technique using the osmium signal from a potassium hexachloroosmate (IV) derivative.