8ooh
From Proteopedia
Cryo-EM map of the focused refinement of the subfamily III haloalkane dehalogenase from Haloferax mediterranei dimer forming hexameric assembly.
Structural highlights
FunctionPublication Abstract from PubMedHaloalkane dehalogenase (HLD) enzymes employ an S(N) 2 nucleophilic substitution mechanism to erase halogen substituents in diverse organohalogen compounds. Subfamily I and II HLDs are well-characterized enzymes, but a mode and purpose of multimerization of subfamily III HLDs are unknown. Here we probe the structural organization of DhmeA, a subfamily III HLD-like enzyme from the archaeon Haloferax mediterranei, by combining cryo-electron microscopy (cryo-EM) and X-ray crystallography. We show that full-length wild-type DhmeA forms diverse quaternary structures, ranging from small oligomers to large supramolecular ring-like assemblies of various sizes and symmetries. We optimized sample preparation steps, enabling three-dimensional reconstructions of an oligomeric species by single-particle cryo-EM. Moreover, we engineered a crystallizable mutant (DhmeA(DeltaGG) ) that provided diffraction-quality crystals. The 3.3 A crystal structure reveals that DhmeA(DeltaGG) forms a ring-like 20-mer structure with outer and inner diameter of ~200 A and ~80 A, respectively. An enzyme homodimer represents a basic repeating building unit of the crystallographic ring. Three assembly interfaces (dimerization, tetramerization and multimerization) were identified to form the supramolecular ring that displays a negatively charged exterior, while its interior part harboring catalytic sites is positively charged. Localization and exposure of catalytic machineries suggest a possible processing of large negatively charged macromolecular substrates. This article is protected by copyright. All rights reserved. Multimeric structure of a subfamily III haloalkane dehalogenase-like enzyme solved by combination of cryo-EM and X-ray crystallography.,Chmelova K, Gao T, Polak M, Schenkmayerova A, Croll TI, Shaikh TR, Skarupova J, Chaloupkova R, Diederichs K, Read RJ, Damborsky J, Novacek J, Marek M Protein Sci. 2023 Aug 13:e4751. doi: 10.1002/pro.4751. PMID:37574754[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
| ||||||||||||||||||
