Heterocyclic and Alkyne Terpenoids by Terpene Synthase-Mediated Biotransformation of Non-Natural Prenyl Diphosphates: Access to New Fragrances and Probes.

Two terpene cyclases were used as biocatalytic tool, namely, limonene synthase from Cannabis sativa (CLS) and 5-epi-aristolochene synthase (TEAS) from Nicotiana tabacum. They showed significant substrate flexibility towards non-natural prenyl diphosphates to form novel terpenoids, including core oxa- and thia-heterocycles and alkyne-modified terpenoids. We elucidated the structures of five novel monoterpene-analogues and a known sesquiterpene-analogue. These results reflected the terpene synthases' ability and promiscuity to broaden the pool of terpenoids with structurally complex analogues. Docking studies highlight an on-off conversion of the unnatural substrates.

Arabidopsis thaliana isoprenyl diphosphate synthases produce the C25 intermediate geranylfarnesyl diphosphate.

Isoprenyl diphosphate synthases (IDSs) catalyze some of the most basic steps in terpene biosynthesis by producing the prenyl diphosphate precursors of each of the various terpenoid classes. Most plants investigated have distinct enzymes that produce the short-chain all-trans (E) prenyl diphosphates geranyl diphosphate (GDP, C10 ), farnesyl diphosphate (FDP, C15 ) or geranylgeranyl diphosphate (GGDP, C20 ). In the genome of Arabidopsis thaliana, 15 trans-product-forming IDSs are present. Ten of these have recently been shown to produce GGDP by genetic complementation of a carotenoid pathway engineered into Escherichia coli. When verifying the product pattern of IDSs producing GGDP by a new LC-MS/MS procedure, we found that five of these IDSs produce geranylfarnesyl diphosphate (GFDP, C25 ) instead of GGDP as their major product in enzyme assays performed in vitro. Over-expression of one of the GFDP synthases in A. thaliana confirmed the production of GFDP in vivo. Enzyme assays with A. thaliana protein extracts from roots but not other organs showed formation of GFDP. Furthermore, GFDP itself was detected in root extracts. Subcellular localization studies in leaves indicated that four of the GFDP synthases were targeted to the plastoglobules of the chloroplast and one was targeted to the mitochondria. Sequence comparison and mutational studies showed that the size of the R group of the 5th amino acid residue N-terminal to the first aspartate-rich motif is responsible for C25 versus C20 product formation, with smaller R groups (Ala and Ser) resulting in GGDP (C20 ) as a product and a larger R group (Met) resulting in GFDP (C25 ).

Similarities and differences in the treatment of spine trauma between surgical specialties and location of practice.

STUDY DESIGN: Questionnaires administered to practicing orthopedic and neurosurgical spine surgeons from various regions of the United States and abroad. OBJECTIVES: To determine similarities and differences in the treatment of spinal trauma. SUMMARY OF BACKGROUND DATA: Spinal trauma is generally referred to subspecialists of orthopedic or neurosurgical training. Prior studies have suggested that there is significant variability in the management of such injuries. METHODS: Questionnaires based on eight clinical scenarios of commonly encountered cervical, thoracic, and lumbar injuries were administered to 35 experienced spinal surgeons. Surgeons completed profile information and answered approximately one dozen questions for each case. Data were analyzed with SPSS software to determine the levels of agreement and characteristics of respondents that might account for a lack of agreement on particular aspects of management. RESULTS: Of the 35 surgeons completing the questionnaire, 63% were orthopedists, 37% were neurosurgeons, and 80% had been in practice for more than 5 years. Considerable agreement was found in the majority of clinical decisions, including whether or not to operate and the timing of surgery. Of the differences noted, neurosurgeons were more likely to obtain a MRI, and orthopedists were more likely to use autograft as a sole graft material. Physicians from abroad were, in general, more likely to operate and to use an anterior approach during surgery than physicians from the northeastern United States. CONCLUSIONS: More commonalities were identified in the management of spinal trauma than previously reported. When found, variability in opinion was related to professional and regional differences.