2PS6

N225D/S229T trichodiene synthase

Summary for 2PS6

• Entry DOI: 10.2210/pdb2ps6/pdb
• Related: 1JFA 1KIY 1YJ4 2AEK 2PS4 2PS7
• Descriptor: Trichodiene synthase, 1,2-ETHANEDIOL (3 entities in total)
• Functional Keywords: terpenoid synthase fold, site-directed mutagenesis, nse/dte motif, magnesium, ethylene glycol, lyase
• Biological source: Fusarium sporotrichioides
• Total number of polymer chains: 2
• Total formula weight: 88255.36

Authors

Vedula, L.S.,Cane, D.E.,Christianson, D.W. (deposition date: 2007-05-04, release date: 2007-12-18, Last modification date: 2023-08-30)

Primary citation

Vedula, L.S.,Jiang, J.,Zakharian, T.,Cane, D.E.,Christianson, D.W.
Structural and mechanistic analysis of trichodiene synthase using site-directed mutagenesis: probing the catalytic function of tyrosine-295 and the asparagine-225/serine-229/glutamate-233-Mg2+B motif.
Arch.Biochem.Biophys., 469:184-194, 2008

PubMed Abstract: Trichodiene synthase from Fusarium sporotrichioides contains two metal ion-binding motifs required for the cyclization of farnesyl diphosphate: the "aspartate-rich" motif D(100)DXX(D/E) that coordinates to Mg2+A and Mg2+C, and the "NSE/DTE" motif N(225)DXXSXXXE that chelates Mg2+B (boldface indicates metal ion ligands). Here, we report steady-state kinetic parameters, product array analyses, and X-ray crystal structures of trichodiene synthase mutants in which the fungal NSE motif is progressively converted into a plant-like DDXXTXXXE motif, resulting in a degradation in both steady-state kinetic parameters and product specificity. Each catalytically active mutant generates a different distribution of sesquiterpene products, and three newly detected sesquiterpenes are identified. In addition, the kinetic and structural properties of the Y295F mutant of trichodiene synthase were found to be similar to those of the wild-type enzyme, thereby ruling out a proposed role for Y295 in catalysis.

PubMed: 17996718
DOI: 10.1016/j.abb.2007.10.015

Experimental method

X-RAY DIFFRACTION (2.6 Å)