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Structural basis for the broad specificity of a new family of amino-acid racemases
Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
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2014 (Engelska)Ingår i: Acta Crystallographica Section D: Biological Crystallography, ISSN 0907-4449, E-ISSN 1399-0047, Vol. 70, s. 79-90Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Broad-spectrum amino-acid racemases (Bsrs) enable bacteria to generate noncanonical D-amino acids, the roles of which in microbial physiology, including the modulation of cell-wall structure and the dissolution of biofilms, are just beginning to be appreciated. Here, extensive crystallographic, mutational, biochemical and bioinformatic studies were used to define the molecular features of the racemase BsrV that enable this enzyme to accommodate more diverse substrates than the related PLP-dependent alanine racemases. Conserved residues were identified that distinguish BsrV and a newly defined family of broad-spectrum racemases from alanine racemases, and these residues were found to be key mediators of the multispecificity of BrsV. Finally, the structural analysis of an additional Bsr that was identified in the bioinformatic analysis confirmed that the distinguishing features of BrsV are conserved among Bsr family members.

Ort, förlag, år, upplaga, sidor
Wiley-Blackwell, 2014. Vol. 70, s. 79-90
Nationell ämneskategori
Biokemi och molekylärbiologi Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci)
Identifikatorer
URN: urn:nbn:se:umu:diva-86070DOI: 10.1107/S1399004713024838ISI: 000329942900010OAI: oai:DiVA.org:umu-86070DiVA, id: diva2:697253
Forskningsfinansiär
Knut och Alice Wallenbergs StiftelseTillgänglig från: 2014-02-17 Skapad: 2014-02-17 Senast uppdaterad: 2019-03-28Bibliografiskt granskad
Ingår i avhandling
1. Uncovering novel cell wall chemistries in gram negative bacteria: from development or dedicated peptidoglycan chemometric tools to functional genomics
Öppna denna publikation i ny flik eller fönster >>Uncovering novel cell wall chemistries in gram negative bacteria: from development or dedicated peptidoglycan chemometric tools to functional genomics
2019 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

Bacteria are surrounded by an external cell wall whose main component is a polymeric net-like structure called the peptidoglycan (PG) or murein sacculus. PG plays crucial roles in bacterial physiology (eg morphogenesis, growth fitness and regulation of innate immunity). Based on the characteristics of this macromolecule, bacteria are grouped as gram negative and positive. Gram negatives present a thin PG layer in the periplasmic space, while Gram positive bacteria contain one thick multi-layered sacculus covering the cytoplasmic membrane. Although the PG sacculus is widely conserved between bacteria, variations in its chemical structure (ie sugars and peptide components) have been reported as a coping mechanism to stress. For example, V. choleraeis able to downregulate PG biosynthesis through non-canonical D-amino acids (NCDAAs) cell wall editing when entering stationary phase. NCDAAs production relies on Bsr enzymes, broad spectrum racemases which are expressed in V. cholerae under the control of stress sigma factor RpoS. In this thesis, we present a comprehensive study that allows us to determine the basic structural and biochemical features required for prominent D-amino acid production by Bsr enzymes.

V. cholerae’s PG editing by NCDAAs revealed the existence of previously unappreciated  chemical modification in the cell wall of bacteria. Such an observation made us question whether the latest technology could reveal, otherwise undetectable, novel PG traits and furthermore, revisit the existence of murein in bacteria which were previously defined as PG-less. Finally, these studies would promote a global assessment of the degree of PG-chemical variability at a Kingdom scale.

On the search for novel functional chemistries and associated mechanisms of cell wall regulation, we analysed the cell wall of hundreds of different species. Here, I present two proof of concept studies: i) investigation of the existence of PG in the Plantomycetes Kuenenia stuttgartiensis, a species previously classified as PG-less; and ii) PG chemical diversity within Class Alphaproteobacteria. To do so, we developed and experimentally validated an innovative chemometric pipeline to rapidly analyse large PG datasets. Chemometric analyses revealed 3 PG clusters within Alphaproteobacteria, which included unprecedented PG modifications widely conserved in family Acetobacteria: amidation at the α-(L)-carboxyl of meso-diaminopimelic acid and the presence of (1–3) cross-linked muropeptides between L-Ala and D-(meso)-diaminopimelate residues from adjacent moieties. Fluctuations of the relative abundance of these PG traits were growth phase and media composition dependent. Functional studies demonstrated that Acetobacteria atypical muropeptides enabled cellular protection against Type VI secreted endopeptidases and negatively affected innate immune system recognition suggesting relevant functional roles in the environmental adaptability of these bacteria.

Ort, förlag, år, upplaga, sidor
Umeå: Umeå University, 2019. s. 63
Serie
Umeå University medical dissertations, ISSN 0346-6612 ; 2027
Nyckelord
Bacteria, cell wall, peptidoglycan, peptidoglycan variations, D-amino acids
Nationell ämneskategori
Mikrobiologi
Forskningsämne
molekylärbiologi
Identifikatorer
urn:nbn:se:umu:diva-157645 (URN)978-91-7855-045-6 (ISBN)
Disputation
2019-04-26, Hörsal d Unod T 9, Umeå University Hospital, Umeå, 09:00 (Engelska)
Opponent
Handledare
Tillgänglig från: 2019-04-05 Skapad: 2019-03-28 Senast uppdaterad: 2019-04-05Bibliografiskt granskad

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Espaillat, AkbarAlvarez, LauraCava, Felipe

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Espaillat, AkbarAlvarez, LauraCava, Felipe
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Institutionen för molekylärbiologi (Medicinska fakulteten)Molekylär Infektionsmedicin, Sverige (MIMS)Umeå Centre for Microbial Research (UCMR)
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Acta Crystallographica Section D: Biological Crystallography
Biokemi och molekylärbiologiMedicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci)

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