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Alanyl‐phosphatidylglycerol synthase: mechanism of substrate recognition during tRNA‐dependent lipid modification in Pseudomonas aeruginosa
Author(s) -
Hebecker Stefanie,
Arendt Wiebke,
Heinemann Ilka U.,
Tiefenau Jana H. J.,
Nimtz Manfred,
Rohde Manfred,
Söll Dieter,
Moser Jürgen
Publication year - 2011
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2011.07621.x
Subject(s) - phosphatidylglycerol , aminoacylation , biochemistry , biology , enzyme , biosynthesis , phospholipid , transfer rna , membrane , rna , gene , phosphatidylcholine
Summary Bacterial lipid homeostasis plays an important role for the adaptation to changing environments and under conditions of antimicrobial treatment. The tRNA‐dependent aminoacylation of the phospholipid phosphatidylglycerol catalysed by aminoacyl‐phosphatidylglycerol synthases was shown to render various organisms less susceptible to antibacterial agents. Therefore, this type of enzyme might provide a new target to potentiate the efficacy of existing antimicrobials. This study makes use of the Pseudomonas aeruginosa alanyl‐phosphatidylglycerol synthase to identify the minimal core domain of this transmembrane protein, which is capable of alanyl‐phosphatidylglycerol biosynthesis. Using this catalytic fragment we established a reliable activity assay that was used to study the enzymatic mechanism by analysing an overall of 33 mutant proteins in vitro . Substrate recognition was analysed by using aminoacylated microhelices as analogues of the natural tRNA substrate. The enzyme even tolerated mutated versions of this minimal substrate, which indicates that neither the intact tRNA, nor the individual sequence of the acceptor stem is a determinant for substrate recognition. Furthermore, the analysis of derivatives of phosphatidylglycerol indicated that the polar head group of the phospholipid is specifically recognized by the enzyme, whereas modification of an individual fatty acid or even the deletion of a single fatty acid did not abolish A‐PG synthesis.