3VS9

Crystal structure of type III PKS ArsC mutant

Summary for 3VS9

• Entry DOI: 10.2210/pdb3vs9/pdb
• Related: 3VS8
• Descriptor: Type III polyketide synthase, SODIUM ION, TETRAETHYLENE GLYCOL, ... (4 entities in total)
• Functional Keywords: thiolase fold, condensing enzyme, transferase
• Biological source: Azotobacter vinelandii
• Total number of polymer chains: 8
• Total formula weight: 363863.17

Authors

Satou, R.,Miyanaga, A.,Ozawa, H.,Funa, N.,Miyazono, K.,Tanokura, M.,Ohnishi, Y.,Horinouchi, S. (deposition date: 2012-04-23, release date: 2013-04-24, Last modification date: 2023-11-08)

Primary citation

Satou, R.,Miyanaga, A.,Ozawa, H.,Funa, N.,Katsuyama, Y.,Miyazono, K.,Tanokura, M.,Ohnishi, Y.,Horinouchi, S.
Structural basis for cyclization specificity of two Azotobacter type III polyketide synthases: a single amino acid substitution reverses their cyclization specificity
J.Biol.Chem., 288:34146-34157, 2013

PubMed Abstract: Type III polyketide synthases (PKSs) show diverse cyclization specificity. We previously characterized two Azotobacter type III PKSs (ArsB and ArsC) with different cyclization specificity. ArsB and ArsC, which share a high sequence identity (71%), produce alkylresorcinols and alkylpyrones through aldol condensation and lactonization of the same polyketomethylene intermediate, respectively. Here we identified a key amino acid residue for the cyclization specificity of each enzyme by site-directed mutagenesis. Trp-281 of ArsB corresponded to Gly-284 of ArsC in the amino acid sequence alignment. The ArsB W281G mutant synthesized alkylpyrone but not alkylresorcinol. In contrast, the ArsC G284W mutant synthesized alkylresorcinol with a small amount of alkylpyrone. These results indicate that this amino acid residue (Trp-281 of ArsB or Gly-284 of ArsC) should occupy a critical position for the cyclization specificity of each enzyme. We then determined crystal structures of the wild-type and G284W ArsC proteins at resolutions of 1.76 and 1.99 Å, respectively. Comparison of these two ArsC structures indicates that the G284W substitution brings a steric wall to the active site cavity, resulting in a significant reduction of the cavity volume. We postulate that the polyketomethylene intermediate can be folded to a suitable form for aldol condensation only in such a relatively narrow cavity of ArsC G284W (and presumably ArsB). This is the first report on the alteration of cyclization specificity from lactonization to aldol condensation for a type III PKS. The ArsC G284W structure is significant as it is the first reported structure of a microbial resorcinol synthase.

PubMed: 24100027
DOI: 10.1074/jbc.M113.487272

Experimental method

X-RAY DIFFRACTION (1.99 Å)