Structure and activity of DmmA, a marine haloalkane dehalogenase

Abstract

DmmA is a haloalkane dehalogenase (HLD) identified and characterized from the metagenomic DNA of a marine microbial consortium. Dehalogenase activity was detected with 1,3-dibromopropane as substrate, with steady-state kinetic parameters typical of HLDs (K m 5 0.24 6 0.05 mM, k cat 5 2.4 6 0.1 s 21). The 2.2-Å crystal structure of DmmA revealed a fold and active site similar to other HLDs, but with a substantially larger active site binding pocket, suggestive of an ability to act on bulky substrates. This enhanced cavity was shown to accept a range of linear and cyclic substrates, suggesting that DmmA will contribute to the expanding industrial applications of HLDs.

Ancestral Haloalkane Dehalogenases Show Robustness and Unique Substrate Specificity

“…We focused on in silico prediction, production, and biochemical characterization of five ancestral enzymes corresponding to different nodes of the HLD‐II phylogenetic tree and representing the ancestors of the thoroughly characterized dehalogenases DbjA, DbeA, DhaA, DmxA, and DmmA . The present‐day enzymes display considerable functional variations even though they are all closely evolutionary related and share similar structural topology, thus providing good models to investigate structural and functional divergence in the HLD‐II subfamily.…”

“…We focusedo ni ns ilico prediction, production,a nd biochemicalc haracterization of five ancestral enzymes correspondingt od ifferent nodes of the HLD-IIp hylogenetic tree and representing the ancestorsofthe thoroughly characterized dehalogenases DbjA, [43,44] DbeA, [45] DhaA, [46] DmxA, [47] and DmmA. [48] The present-daye nzymes display considerable functional variations even thought hey are all closely evolutionary relateda nd share similars tructural topology,t hus providing good modelst oi nvestigate structural andf unctional divergence in the HLD-II subfamily.Characterization of the resurrected ancestral enzymes revealed unique functional properties,including enhanced thermostability,i mproved specific activity,o r modified substrate specificity.T he reconstructed enzymes are the most thermodynamically stable HLDs. Our study highlights the benefits of ASRf or developing novel biocatalysts.…”

A mini review of haloalkane dehalogenase: From molecular characterization to applications

“…To date, several native structures of HLDs have been published and made available in Protein Data Bank. They include DhlA isolated from X. authotropicus GJ10 [9], DhaA from R. rhodochrous NCIMB13064 [5], LinB from S. paucimobilis UT26 [2], DmbA from M. tuberculosis Rv2579 [10], DbjA from B. japonicum USDA110 [11], DppA from P. pacifica SIR-1 [12], DmmA from M. producta [13] and DatA from Agrobacterium tumefaciens C58 [14].…”

“…In general, structure of HLDs consists of main domain with an eight-stranded parallel -sheet structure and; connected by loops; -helices cap domain on top of main domain. The main domain and cap domain form a strong internal hydrophobic cavity which is deeply buried inside the core of the enzyme [2,5,[9][10][12][13][14][15][16]. The /-hydrolase domain fold are the most frequently found in nature and are adopted by largest groups of proteins that display an enormous diversity in sequence, fold plasticity and catalytic functions and, moreover, in the evolution point of view, this fold diverge from a common ancestor [17][18][19][20].…”

“…They play significant role in the halide binding; via hydrogen bond; and maintain nucleophile in proper orientation, usually serves by highly conserved hydrophobic residue i.e. tryptophan [12][13]27,31]. Variability of first halide stabilizing residue was firstly observed in HLD, DatA, from A. tumefaciens C58.…”