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Synthetic Phosphodiester‐Linked 4‐Amino‐4‐deoxy‐ l ‐arabinose Derivatives Demonstrate that ArnT is an Inverting Aminoarabinosyl Transferase
Author(s) -
Olag Charlotte,
Monjaras Feria Julia,
GrünwaldGruber Clemens,
Blaukopf Markus,
Valvano Miguel A.,
Kosma Paul
Publication year - 2019
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201900349
Subject(s) - chemistry , transferase , lipid a , stereochemistry , phosphodiester bond , anomer , aquifex aeolicus , biochemistry , escherichia coli , enzyme , lipopolysaccharide , gene , rna , medicine , endocrinology
4‐Amino‐4‐deoxy‐ l ‐arabinopyranose (Ara4N) residues have been linked to antibiotic resistance due to reduction of the negative charge in the lipid A and core regions of the bacterial lipopolysaccharide (LPS). To study the enzymatic transfer of Ara4N onto lipid A, which is catalysed by the ArnT transferase, we chemically synthesised a series of anomeric phosphodiester‐linked lipid Ara4N derivatives containing linear aliphatic chains as well as E ‐ and Z ‐configured monoterpene units. Coupling reactions were based on sugar‐derived H ‐phosphonates, followed by oxidation and global deprotection. The enzymatic Ara4N transfer was performed in vitro with crude membranes from a deep‐rough mutant from Escherichia coli as acceptor. Product formation was detected by TLC and LC‐ESI‐QTOF mass spectrometry. Out of seven analogues tested, only the α‐neryl derivative was accepted by the Burkholderia cenocepacia ArnT protein, leading to substitution of the Kdo 2 ‐lipid A acceptor and thus affording evidence that ArnT is an inverting glycosyl transferase that requires the Z ‐configured double bond next to the anomeric phosphate moiety. This approach provides an easily accessible donor substrate for biochemical studies relating to modifications of bacterial LPS that modulate antibiotic resistance and immune recognition.