Biosynthesis of Streptolidine Involved Two Unexpected Intermediates Produced by a Dihydroxylase and a Cyclase through Unusual Mechanisms |
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| Authors: | Dr. Chin‐Yuan Chang Syue‐Yi Lyu Dr. Yu‐Chen Liu Ning‐Shian Hsu Dr. Chih‐Chung Wu Dr. Cheng‐Fong Tang Kuan‐Hung Lin Dr. Jin‐Yuan Ho Dr. Chang‐Jer Wu Dr. Ming‐Daw Tsai Dr. Tsung‐Lin Li |
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| Affiliation: | 1. Genomics Research Center, Academia Sinica, 128 Academia Road, Section 2, Nankang, Taipei 115 (Taiwan);2. Institute of Biological Chemistry, Academia Sinica, 128 Academia Road, Section 2, Nankang, Taipei 115 (Taiwan);3. Department of Food Science, National Taiwan Ocean University, 2 Pei‐Ning Road, Keeling 20224 (Taiwan);4. Biotechnology Center, National Chung Hsing University, 250 Kuo Kuang Road, Taichung 402 (Taiwan) |
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| Abstract: | Streptothricin‐F (STT‐F), one of the early‐discovered antibiotics, consists of three components, a β‐lysine homopolymer, an aminosugar D ‐gulosamine, and an unusual bicyclic streptolidine. The biosynthesis of streptolidine is a long‐lasting but unresolved puzzle. Herein, a combination of genetic/biochemical/structural approaches was used to unravel this problem. The STT gene cluster was first sequenced from a Streptomyces variant BCRC 12163, wherein two gene products OrfP and OrfR were characterized in vitro to be a dihydroxylase and a cyclase, respectively. Thirteen high‐resolution crystal structures for both enzymes in different reaction intermediate states were snapshotted to help elucidate their catalytic mechanisms. OrfP catalyzes an FeII‐dependent double hydroxylation reaction converting L ‐Arg into (3R,4R)‐(OH)2‐L ‐Arg via (3S)‐OH‐L ‐Arg, while OrfR catalyzes an unusual PLP‐dependent elimination/addition reaction cyclizing (3R,4R)‐(OH)2‐L ‐Arg to the six‐membered (4R)‐OH‐capreomycidine. The biosynthetic mystery finally comes to light as the latter product was incorporation into STT‐F by a feeding experiment. |
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| Keywords: | biosynthesis enzymes isotopic labeling protein structures reaction mechanisms |
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