(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > other sequences: NE > artificial sequences: NE > synthetic construct: NE
Haloalkane_dehalogenase-HLD2 : 9zzzz-AncFT7 Synthetic construct of Renilla-type engineered ancestral luciferase variant (AncFT7), 9zzzz-AncHLDRLuc2 Synthetic construct of the common ancestor of haloalkane dehalogenases and Renilla luciferase with fragment transplabntation (Anc-FT) AncFT, 9zzzz-AncHLDRLuc Synthetic construct of the common ancestor of haloalkane dehalogenases and Renilla luciferase. Hydroxynitrile_lyase : 9zzzz-5TDX Resurrected ancestral hydroxynitrile lyase from flowering plants
Molecular evidence
Database
No mutation 1 structure: 7PW1: Crystal structure of ancestral haloalkane dehalogenase AncLinB-DmbA No kinetic
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA MTALGAEPYGQKKFIEIAGKRMAYIDEGEGDPIVFQHGNPTSSYLWRNIM PHLEGLGRLIACDLIGMGDSDKLSPSGPDRYSYAEHRDYLFALWEALDLG DNVVLVIHDWGSALGFDWANQHRDRVQGIAYMEAIVTPLEWADWPEEVRD IFQGFRSPAGEEMVLENNIFVERVLPGAILRQLSDEEMAEYRRPFLNAGE DRRPTLSWPRQIPIDGEPADVVAIVSDYASWLAESDIPKLFINAEPGAIV TGRMRDFCRSWPNQTEITVKGAHFIQEDSPDEIGAAIAEFVRRLRAAAGV
Haloalkane dehalogenases (EC 3.8.1.5) play an important role in hydrolytic degradation of halogenated compounds, resulting in a halide ion, a proton, and an alcohol. They are used in biocatalysis, bioremediation, and biosensing of environmental pollutants and also for molecular tagging in cell biology. The method of ancestral sequence reconstruction leads to prediction of sequences of ancestral enzymes allowing their experimental characterization. Based on the sequences of modern haloalkane dehalogenases from the subfamily II, the most common ancestor of thoroughly characterized enzymes LinB from Sphingobium japonicum UT26 and DmbA from Mycobacterium bovis 5033/66 was in silico predicted, recombinantly produced and structurally characterized. The ancestral enzyme AncLinB-DmbA was crystallized using the sitting-drop vapor-diffusion method, yielding rod-like crystals that diffracted X-rays to 1.5 A resolution. Structural comparison of AncLinB-DmbA with their closely related descendants LinB and DmbA revealed some differences in overall structure and tunnel architecture. Newly prepared AncLinB-DmbA has the highest active site cavity volume and the biggest entrance radius on the main tunnel in comparison to descendant enzymes. Ancestral sequence reconstruction is a powerful technique to study molecular evolution and design robust proteins for enzyme technologies.
Ancestral Haloalkane Dehalogenase AncLinB-DmbA
