β-alanine synthase : one reaction, two folds and mechanisms

<p>beta-alanine synthase (betaAS) is the third enzyme in the reductive pyrimidine catabolic pathway which is responsible for the breakdown of pyrimidine bases, including several anti-cancer drugs in higher organisms. We have solved the high resolution structures of two beta-alanine synthases that perform the same reaction but have two very different folds and utilize different reaction mechanisms.</p><p>The fold of the homodimeric betaAS from the yeast Saccharomyces kluyveri (SkbetaAS) identifies it as a member of the Acy1/M20 family of metallopeptidases. Its subunit consists of two domains, of which the larger harbors a di-zinc centre crucial for catalysis, while the smaller domain mediates dimerization. We determined the structure of a productive substrate complex, the first for a dimeric member of the Acy1/M20 family. With this structure we show that a conformational change from an open state to a closed state is required for catalysis. Additionally the results of our site-directed mutagenesis studies, performed to identify residues involved in substrate binding and catalysis are discussed. These results support the role of E159 as a catalytic base and identify R322 as a key substrate-binding residue.</p><p>We have also characterized the structure of betaAS from Drosophila melanogaster (DmbetaAS) which has been identified as a member of the nitrilase superfamily, with a characteristic alphabetabetaalpha-sandwich fold and a conserved Cys-Glu-Lys catalytic triad. We have determined the threedimensional structure in two different space groups, in P21212 to 2.8 Å and in C2 to 3.3 Å resolution, respectively. The first crystal form has four molecules in the asymmetric unit, forming a homotetramer in a dimer-ofdimers arrangement. This homotetramer is assembled into an octamer with the molecule from an adjacent asymmetric unit in a helical turn-like assembly. The same octameric formation is found in the second crystal form, which has all eight monomers present in the asymmetric unit.</p><h3>List of scientific papers</h3><p>I. Lundgren S, Gojkoviæ Z, Piskur J, Dobritzsch D (2003). "Yeast beta-alanine synthase shares a structural scaffold and origin with dizinc-dependent exopeptidases." J Biol Chem 278(51): 51851-62. Epub 2003 Oct 8 <br><a href="https://pubmed.ncbi.nlm.nih.gov/14534321">https://pubmed.ncbi.nlm.nih.gov/14534321</a><br><br></p><p>II. Lundgren S, Andersen B, Piskur J, Dobritzsch D (2007). "Crystal structures of yeast -alanine synthase complexes reveal the mode of substrate binding and large scale domain closure movements." J Biol Chem 282(49): 36037-47. Epub 2007 Oct 4 <br><a href="https://pubmed.ncbi.nlm.nih.gov/17916556">https://pubmed.ncbi.nlm.nih.gov/17916556</a><br><br></p><p>III. Lundgren S, Andersen B, Piskur J, Dobritzsch D (2007). "Crystallization and preliminary X-ray data analysis of beta-alanine synthase from Drosophila melanogaster." Acta Crystallogr Sect F Struct Biol Cryst Commun 63(Pt 10): 874-7. Epub 2007 Sep 19 <br><a href="https://pubmed.ncbi.nlm.nih.gov/17909293">https://pubmed.ncbi.nlm.nih.gov/17909293</a><br><br></p><p>IV. Lundgren S, Lohkamp B, Andersen B, Piskur J, Dobritzsch D (2007). "The crystal structure of beta-alanine synthase from Drosophila melanogaster reveals a nitrilase fold and an octameric helical turnlike assembly." J Mol Biol (Submitted)</p>