Abstract
Background: FabB (3-oxoacyl-[acyl-carrier-protein] synthase 1) is part of the fatty acid synthesis II pathway found in bacteria and a potential target for antibiotics. The enzyme catalyses the Claisen condensation of malonyl-ACP (acyl carrier protein) with acyl-ACP via an acyl-enzyme intermediate. Here, we report the crystal structure of the intermediate-mimicking Pseudomonas aeruginosa FabB (PaFabB) C161A variant. Methods: His-tagged PaFabB C161A was expressed in E. coli Rosetta DE3 pLysS cells, cleaved by TEV protease and purified using affinity and size exclusion chromatography. Commercial screens were used to identify suitable crystallization conditions which were subsequently improved to obtain well diffracting crystals. Results: We developed a robust and efficient system for recombinant expression of PaFabB C161A. Conditions to obtain well diffracting crystals were established. The crystal structure of PaFabB C161A was solved by molecular replacement at 1.3 Å resolution. Binding site comparison between PaFabB and PaFabF revealed a conserved malonyl binding site but differences in the fatty acid binding channel. Conclusions: The PaFabB C161A crystal structure can be used as a template to facilitate the design of FabB inhibitors.
Highlights
New antibiotics are urgently needed to maintain the high standard of living that we have got accustomed to as the antibiotics of today are losing effectiveness faster than they are being replaced by new treatment options.[1]
Some figures illustrating the structure of Pseudomonas aeruginosa FabB (PaFabB) C161A have been combined and revised to make the article easier to comprehend and less repetitive
Genes coding for enzymes in the FAS II pathway have been found to be essential for P. aeruginosa in several genetic screens, including the gene for FabB (3-oxoacyl-[acyl-carrier-protein] synthase 1).[4,5,6,7,8]
Summary
Any reports and responses or comments on the article can be found at the end of the article. This article is included in the Chemical Information Science gateway. A comparison of the fatty acid binding channel in PaFabB and PaFabF has been added. Some figures illustrating the structure of PaFabB C161A have been combined and revised to make the article easier to comprehend and less repetitive. The role of an active site Phe in substrate and ligand binding and how active site mutations influence the conformation of this residue have been explained with more detail. We have made some minor changes to the text as suggested by the reviewers. Any further responses from the reviewers can be found at the end of the article
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