期刊
ACS CHEMICAL BIOLOGY
卷 15, 期 6, 页码 1517-1525出版社
AMER CHEMICAL SOC
DOI: 10.1021/acschembio.0c00145
关键词
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资金
- JSPS KAKENHI [17H05439, 19H04652, 18H02145, 19K22273, 19H04648, 19H04661]
- Takeda Science Foundation
- Platform Project for Supporting in Drug Discovery and Life Science Research from Japan Agency for Medical Research and Development (AMED) [BINDS1447, 2018RP-27, 2019RP-31]
- Grants-in-Aid for Scientific Research [19H04648, 19H04652, 19H04661, 19K22273, 18H02145, 17H05439] Funding Source: KAKEN
Terpene synthases (TS) are classified into two broad types, Class I and II, based on the chemical strategy for initial carbocation formation and motif sequences of the catalytic site. We have recently identified a new class of enzymes, Class IB, showing the acceptability of long (C-20-C-35) prenyl-diphosphates as substrates and no amino acid sequence homology with known TS. Conversion of long prenyl-diphosphates such as heptaprenyl-diphosphate (C-35) is unusual and has never been reported for Class I and II enzymes. Therefore, the characterization of Class IB enzymes is crucial to understand the reaction mechanism of the extensive terpene synthesis. Here, we report the crystal structure bound with a substrate surrogate and biochemical analysis of a Class IB TS, using the enzyme from Bacillus alcalophilus (BaITS). The structure analysis revealed that the diphosphate part of the substrate is located around the two characteristic Asp-rich motifs, and the hydrophobic tail is accommodated in a unique hydrophobic long tunnel, where the C-35 prenyl-diphosphate, the longest substrate of BalTS, can be accepted. Biochemical analyses of BalTS showed that the enzymatic property, such as Mg2+ dependency, is similar to those of Class I enzymes. In addition, a new cyclic terpene was identified from BaITS reaction products. Mutational analysis revealed that five of the six Asp residues in the Asp-rich motifs and two His residues are essential for the formation of the cydic skeleton. These results provided a clue to consider the application of the unusual large terpene synthesis by Class IB enzymes.
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