4HHQ

Serum paraoxonase-1 by directed evolution with the H115Q and H134Q mutations

Summary for 4HHQ

• Entry DOI: 10.2210/pdb4hhq/pdb
• Related: 1V04 3SRE 3SRG 4HHO
• Descriptor: SERUM PARAOXONASE BY DIRECTED EVOLUTION, CALCIUM ION, DODECYL-BETA-D-MALTOSIDE, ... (5 entities in total)
• Functional Keywords: 6-blades -propeller fold, hydrolase
• Biological source: synthetic construct (artificial gene)
• Total number of polymer chains: 1
• Total formula weight: 40414.40

Authors

Moshe, B.-D.,Grzegorz, W.,Mikael, E.,Israel, S.,Joel, L.S.,Dan, S.T. (deposition date: 2012-10-10, release date: 2013-03-27, Last modification date: 2024-11-20)

Primary citation

Ben-David, M.,Wieczorek, G.,Elias, M.,Silman, I.,Sussman, J.L.,Tawfik, D.S.
Catalytic metal ion rearrangements underline promiscuity and evolvability of a metalloenzyme.
J.Mol.Biol., 425:1028-1038, 2013

PubMed Abstract: Although largely deemed as structurally conserved, catalytic metal ion sites can rearrange, thereby contributing to enzyme evolvability. Here, we show that in paraoxonase-1, a lipo-lactonase, catalytic promiscuity and divergence into an organophosphate hydrolase are correlated with an alternative mode of the catalytic Ca(2+). We describe the crystal structures of active-site mutants bearing mutations at position 115. The histidine at this position acts as a base to activate the lactone-hydrolyzing water molecule. Mutations to Trp or Gln indeed diminish paraoxonase-1's lactonase activity; however, the promiscuous organophosphate hydrolase activity is enhanced. The structures reveal a 1.8-Å upward displacement towards the enzyme's surface of the catalytic Ca(2+) in the His115 mutants and configurational changes in the ligating side chains and water molecules, relative to the wild-type enzyme. Biochemical analysis and molecular dynamics simulations suggest that this alternative, upward metal mode mediates the promiscuous hydrolysis of organophosphates. The upward Ca(2+) mode observed in the His115 mutants also appears to mediate the wild type's paraoxonase activity. However, whereas the upward mode dominates in the Trp115 mutant, it is scarcely populated in wild type. Thus, the plasticity of active-site metal ions may permit alternative, latent, promiscuous activities and also provide the basis for the divergence of new enzymatic functions.

PubMed: 23318950
DOI: 10.1016/j.jmb.2013.01.009

Experimental method

X-RAY DIFFRACTION (2.3 Å)