title: Characterization of serine hydroxymethyltransferase GlyA as a potential source of D-alanine in Chlamydia pneumoniae.
abstract: For intracellular Chlamydiaceae, there is no need to withstand osmotic challenges, and a functional cell wall has not been detected in these pathogens so far. Nevertheless, penicillin inhibits cell division in Chlamydiaceae resulting in enlarged aberrant bodies, a phenomenon known as chlamydial anomaly. D-alanine is a unique and essential component in the biosynthesis of bacterial cell walls. In free-living bacteria like Escherichia coli, penicillin-binding proteins such as monofunctional transpeptidases PBP2 and PBP3, the putative targets of penicillin in Chlamydiaceae, cross-link adjacent peptidoglycan strands via meso-diaminopimelic acid and D-Ala-D-Ala moieties of pentapeptide side chains. In the absence of genes coding for alanine racemase Alr and DadX homologs, the source of D-Ala and thus the presence of substrates for PBP2 and PBP3 activity in Chlamydiaceae has puzzled researchers for years. Interestingly, Chlamydiaceae genomes encode GlyA, a serine hydroxymethyltransferase that has been shown to exhibit slow racemization of D- and L-alanine as a side reaction in E. coli. We show that GlyA from Chlamydia pneumoniae can serve as a source of D-Ala. GlyA partially reversed the D-Ala auxotrophic phenotype of an E. coli racemase double mutant. Moreover, purified chlamydial GlyA had racemase activity on L-Ala in vitro and was inhibited by D-cycloserine, identifying GlyA, besides D-Ala ligase MurC/Ddl, as an additional target of this competitive inhibitor in Chlamydiaceae. Proof of D-Ala biosynthesis in Chlamydiaceae helps to clarify the structure of cell wall precursor lipid II and the role of chlamydial penicillin-binding proteins in the development of non-dividing aberrant chlamydial bodies and persistence in the presence of penicillin.
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Stefania
authors.initials: S
authors.lastname: De Benedetti
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Henrike
authors.initials: H
authors.lastname: Bühl
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Gaballah
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Anna
authors.initials: A
authors.lastname: Klöckner
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Otten
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Tanja
authors.initials: T
authors.lastname: Schneider
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Hans-Georg
authors.initials: HG
authors.lastname: Sahl
authors.affiliation: Pharmaceutical Microbiology Section, Institute for Medical Microbiology, Immunology and Parasitology, University of Bonn Bonn, Germany.
authors.forename: Beate
authors.initials: B
authors.lastname: Henrichfreise
mesh_terms: D000409:Alanine / Q000378:metabolism*; D000411:Alanine Racemase / Q000235:genetics / Q000378:metabolism*; D016993:Chlamydophila pneumoniae / Q000201:enzymology* / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D012696:Glycine Hydroxymethyltransferase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D012696:Glycine Hydroxymethyltransferase; D000411:Alanine Racemase; D000409:Alanine
keywords: D-alanine; D-cycloserine; GlyA; aberrant bodies; alanine racemase; chlamydial anomaly; penicillin; persistence