title: The crystal structure of D-threonine aldolase from Alcaligenes xylosoxidans provides insight into a metal ion assisted PLP-dependent mechanism.
abstract: Threonine aldolases catalyze the pyridoxal phosphate (PLP) dependent cleavage of threonine into glycine and acetaldehyde and play a major role in the degradation of this amino acid. In nature, L- as well as D-specific enzymes have been identified, but the exact physiological function of D-threonine aldolases (DTAs) is still largely unknown. Both types of enantio-complementary enzymes have a considerable potential in biocatalysis for the stereospecific synthesis of various β-hydroxy amino acids, which are valuable building blocks for the production of pharmaceuticals. While several structures of L-threonine aldolases (LTAs) have already been determined, no structure of a DTA is available to date. Here, we report on the determination of the crystal structure of the DTA from Alcaligenes xylosoxidans (AxDTA) at 1.5 Å resolution. Our results underline the close relationship of DTAs and alanine racemases and allow the identification of a metal binding site close to the PLP-cofactor in the active site of the enzyme which is consistent with the previous observation that divalent cations are essential for DTA activity. Modeling of AxDTA substrate complexes provides a rationale for this metal dependence and indicates that binding of the β-hydroxy group of the substrate to the metal ion very likely activates this group and facilitates its deprotonation by His193. An equivalent involvement of a metal ion has been implicated in the mechanism of a serine dehydratase, which harbors a metal ion binding site in the vicinity of the PLP cofactor at the same position as in DTA. The structure of AxDTA is completely different to available structures of LTAs. The enantio-complementarity of DTAs and LTAs can be explained by an approximate mirror symmetry of crucial active site residues relative to the PLP-cofactor.
authors.affiliation: Austrian Centre of Industrial Biotechnology, Petersgasse 14, 8010, Graz, Austria.
authors.forename: Michael K
authors.initials: MK
authors.lastname: Uhl
authors.affiliation: Austrian Centre of Industrial Biotechnology, Petersgasse 14, 8010, Graz, Austria; Institute of Molecular Biosciences, University of Graz, Humboldtstraße 50/3, 8010, Graz, Austria.
authors.forename: Gustav
authors.initials: G
authors.lastname: Oberdorfer
authors.affiliation: Austrian Centre of Industrial Biotechnology, Petersgasse 14, 8010, Graz, Austria.
authors.forename: Georg
authors.initials: G
authors.lastname: Steinkellner
authors.affiliation: Institute of Molecular Biosciences, University of Graz, Humboldtstraße 50/3, 8010, Graz, Austria.
authors.forename: Lina
authors.initials: L
authors.lastname: Riegler-Berket
authors.affiliation: DSM Chemical Technology R&D BV - Innovative Synthesis, 6167, Geleen, The Netherlands.
authors.forename: Daniel
authors.initials: D
authors.lastname: Mink
authors.affiliation: DSM Chemical Technology R&D BV - Innovative Synthesis, 6167, Geleen, The Netherlands.
authors.forename: Friso
authors.initials: F
authors.lastname: van Assema
authors.affiliation: DSM Chemical Technology R&D BV - Innovative Synthesis, 6167, Geleen, The Netherlands.
authors.forename: Martin
authors.initials: M
authors.lastname: Schürmann
authors.affiliation: Austrian Centre of Industrial Biotechnology, Petersgasse 14, 8010, Graz, Austria; Institute of Molecular Biosciences, University of Graz, Humboldtstraße 50/3, 8010, Graz, Austria.
authors.forename: Karl
authors.initials: K
authors.lastname: Gruber
mesh_terms: D000079:Acetaldehyde / Q000378:metabolism; D000411:Alanine Racemase / Q000737:chemistry / Q000235:genetics; D000421:Alcaligenes / Q000201:enzymology* / Q000235:genetics; D000446:Aldehyde-Lyases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli; D005998:Glycine / Q000096:biosynthesis; D008345:Manganese / Q000378:metabolism; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D005810:Multigene Family; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011522:Protons; D011732:Pyridoxal Phosphate / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D012694:Serine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D013329:Structure-Activity Relationship; D013912:Threonine / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D011522:Protons; D011994:Recombinant Proteins; D013912:Threonine; D008345:Manganese; D012694:Serine; D011732:Pyridoxal Phosphate; D000446:Aldehyde-Lyases; C108811:threonine acetaldehyde-lyase; D000411:Alanine Racemase; D000079:Acetaldehyde; C467259:phenylserine; D005998:Glycine