6f54

CRYSTAL STRUCTURE OF KETOSTEROID ISOMERASE TRIPLE VARIANT V88I/L99VD103SCRYSTAL STRUCTURE OF KETOSTEROID ISOMERASE TRIPLE VARIANT V88I/L99VD103S

Structural highlights 6f54 is a 2 chain structure with sequence from Pseudomonas putida. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 1.08Å Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function SDIS_PSEPU Publication Abstract from PubMed The realization of a synthetic biology approach to microbial (1R,2S,5R)-(-)-menthol (1) production relies on the identification of a gene encoding an isopulegone isomerase (IPGI), the only enzyme in the Mentha piperita biosynthetic pathway as yet unidentified. We demonstrate that Delta5-3-ketosteroid isomerase (KSI) from Pseudomonas putida can act as an IPGI, producing (R)-(+)-pulegone ((R)-2) from (+)-cis-isopulegone (3). Using a robotics-driven semirational design strategy, we identified a key KSI variant encoding four active site mutations, which confer a 4.3-fold increase in activity over the wild-type enzyme. This was assisted by the generation of crystal structures of four KSI variants, combined with molecular modeling of 3 binding to identify key active site residue targets. The KSI variant was demonstrated to function efficiently within cascade biocatalytic reactions with downstream Mentha enzymes pulegone reductase and (-)-menthone:(-)-menthol reductase to generate 1 from 3. This study introduces the use of a recombinant IPGI, engineered to function efficiently within a biosynthetic pathway for the production of 1 in microorganisms. Engineering the "Missing Link" in Biosynthetic (-)-Menthol Production: Bacterial Isopulegone Isomerase.,Currin A, Dunstan MS, Johannissen LO, Hollywood KA, Vinaixa M, Jervis AJ, Swainston N, Rattray NJW, Gardiner JM, Kell DB, Takano E, Toogood HS, Scrutton NS ACS Catal. 2018 Mar 2;8(3):2012-2020. doi: 10.1021/acscatal.7b04115. Epub 2018, Jan 24. PMID:29750129 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See Also Ketosteroid Isomerase

spinqualitylabelsall models 6f54, resolution 1.08Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image