PDBsum entry: 1u0u

Transferase

PDB id: 1u0u

Contents

Protein chains

(+ 0 more) 389 a.a. *

Waters ×1871

* Residue conservation analysis

PDB id:

1u0u

Name:

Transferase

Title:

An aldol switch discovered in stilbene synthases mediates cyclization specificity of type iii polyketide synthases: pine stilbene synthase structure

Structure:

Dihydropinosylvin synthase. Chain: a, b, c, d, e, f. Synonym: stilbene synthase, sts, pinosylvin-forming stilbene synthase. Engineered: yes

Source:

Pinus sylvestris. Scots pine. Organism_taxid: 3349. Gene: pss1. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Biol. unit:

Dimer (from PQS)

Resolution:

2.11Å R-factor: 0.225 R-free: 0.289

Authors:

M.B.Austin,M.E.Bowman,J.-L.Ferrer,J.Schroder,J.P.Noel

Key ref:

M.B.Austin et al. (2004). An aldol switch discovered in stilbene synthases mediates cyclization specificity of type III polyketide synthases. Chem Biol, 11, 1179-1194. PubMed id: 15380179 DOI: 10.1016/j.chembiol.2004.05.024

Date:

14-Jul-04 Release date: 12-Oct-04

Protein chains

Q02323  (DPSS_PINSY) -  Dihydropinosylvin synthase

Seq: 393 a.a.

Struc: 389 a.a.

 Gene Ontology (GO) functional annotation 

• Cellular component: cytoplasm (1 term)
• Biological process: metabolic process (3 terms)
• Biochemical function: catalytic activity (4 terms)

DOI no: 10.1016/j.chembiol.2004.05.024

Chem Biol 11:1179-1194 (2004)

PubMed id: 15380179

An aldol switch discovered in stilbene synthases mediates cyclization specificity of type III polyketide synthases.

M.B.Austin, M.E.Bowman, J.L.Ferrer, J.Schröder, J.P.Noel.

ABSTRACT

Stilbene synthase (STS) and chalcone synthase (CHS) each catalyze the formation of a tetraketide intermediate from a CoA-tethered phenylpropanoid starter and three molecules of malonyl-CoA, but use different cyclization mechanisms to produce distinct chemical scaffolds for a variety of plant natural products. Here we present the first STS crystal structure and identify, by mutagenic conversion of alfalfa CHS into a functional stilbene synthase, the structural basis for the evolution of STS cyclization specificity in type III polyketide synthase (PKS) enzymes. Additional mutagenesis and enzymatic characterization confirms that electronic effects rather than steric factors balance competing cyclization specificities in CHS and STS. Finally, we discuss the problematic in vitro reconstitution of plant stilbenecarboxylate pathways, using insights from existing biomimetic polyketide cyclization studies to generate a novel mechanistic hypothesis to explain stilbenecarboxylate biosynthesis.

Selected figure(s)

Figure 3.
Figure 3. Thioesterase-like STS “Aldol Switch” Controls Cyclization Specificity(A) Slightly different bound conformations of resveratrol observed in the complexed 18xCHS crystal structure (green and rose) correlate to movements of the flexible Phe265 side chain, overlaid with the structure of the previously determined resveratrol [12] bound in wild-type CHS (light gray) and viewed down the CoA binding tunnel into the active site cavity. Positioning of resveratrol's starter- and malonyl-derived aromatic rings are similar to each other and to CHS-bound naringenin (shown in Figure 1B in a similar view).(B) C-α trace overlay of the displacement of the area 2 loop in STS (gold) and 18xCHS (green), compared to CHS (blue). Two orientations illustrate the positions and movements of residues 131–133 (CHS numbering).(C) Stereoview of the 18xCHS STS-like “aldol switch” hydrogen bonds, showing the 1.9 Å resolution 2F[o] − F[c] electron density map (blue wirecage) contoured at 1 sigma.(D) “Aldol switch” hydrogen bonding differences (resulting from repositioning of the Thr132 side chain) in CHS-like and STS-like active sites, compared to each other and to the active site of thioesterase II (TEII) from E. coli ([18]; PDB code 1C8U). Distances incompatible with hydrogen bond formation are given in parentheses and indicated with double-headed arrows. Putative nucleophilic water positions are highlighted in yellow.(E) Thin layer chromatography (TLC) analysis of the cyclization specificities of mutants designed to disrupt the 18xCHS mutant's aldol switch hydrogen bond network while preserving the 18xCHS STS-like conformational changes (see text).

Figure 4.
Figure 4. STS Mechanistic Options and Relevant Solution Chemistry(A) Spontaneous solution-based polyketide C2→C7 aldol condensation cyclization chemistry leading to stilbenes. Atoms fated for elimination as molecules of CO[2] and H[2]O are colored in red and blue, respectively. The aromatized stilbene acid solution-based intermediate product has been shown not to be an intermediate in the STS-catalyzed reaction (see text).(B) Plausible reaction pathways for the four STS cyclization-related events, assuming mechanistic divergence from CHS begins with an aldol switch-catalyzed thioesterase-like hydrolytic step. Scenario One depicts a decarboxylative cyclization reaction, as described by Ebizuka's group [14]. Scenario Two depicts two alternative decarboxylation schemes that follow a solution chemistry-like nondecarboxylative aldol condensation-based cyclization. Atoms fated for elimination as molecules of CO[2] and H[2]O are again colored red and blue, respectively.

The above figures are reprinted by permission from Cell Press: Chem Biol (2004, 11, 1179-1194) copyright 2004.

Figures were selected by an automated process.