Crystal structure of alkyl hydroperoxidase D like protein PA0269 from Pseudomonas aeruginosa: Homology of the AhpD-like structural family

Abstract

BackgroundAlkyl hydroperoxidase activity provides an important antioxidant defense for bacterial cells. The catalytic mechanism requires two peroxidases, AhpC and AhpD, where AhpD plays the role of an essential adaptor protein.ResultsThe crystal structure of a putative AhpD from Pseudomonas aeruginosa has been determined at 1.9 Å. The protein has an all-helical fold with a chain topology similar to a known AhpD from Mycobacterium tuberculosis despite a low overall sequence identity of 9%. A conserved two α-helical motif responsible for function is present in both. However, in the P. aeruginosa protein, helices H3, H4 of this motif are located at the N-terminal part of the chain, while in M. tuberculosis AhpD, the corresponding helices H8, H9 are situated at the C-terminus. Residues 24-62 of the putative catalytic region of P. aeruginosa have a higher sequence identity of 33% where the functional activity is supplied by a proton relay system of five residues, Glu36, Cys48, Tyr50, Cys51, and His55, and one structural water molecule. A comparison of five other related hypothetical proteins from various species, assigned to the alkyl hydroperoxidase D-like protein family, shows they contain the same conserved structural motif and catalytic sequence Cys-X-X-Cys. We have shown that AhpD from P. aeruginosa exhibits a weak ability to reduce H2O2 as tested using a ferrous oxidation-xylenol orange (FOX) assay, and this activity is blocked by thiol alkylating reagents.ConclusionThus, this hypothetical protein was assigned to the AhpD-like protein family with peroxidase-related activity. The functional relationship of specific oligomeric structures of AhpD-like structural family is discussed.

Janus-Faced Sestrin2 Controls ROS and MTOR Signalling Through Two Separate Functional Domains

“…Despite barely conserved primary sequences ( Supplementary Fig. 5b), we noted that 109-139 amino acids of the Sesn-A domain show a very distant sequence homology to YP_296737.1 as well as to AhpD, a well-characterized alkylhydroperoxidase in Mycobacterium tuberculosis [21][22][23] , as formerly reported 2 . The homology region corresponds to the helix-turn-helix motif of AhpD, a signature motif found in the family of AhpD-like oxidoreductases [21][22][23] .…”

“…5b), we noted that 109-139 amino acids of the Sesn-A domain show a very distant sequence homology to YP_296737.1 as well as to AhpD, a well-characterized alkylhydroperoxidase in Mycobacterium tuberculosis [21][22][23] , as formerly reported 2 . The homology region corresponds to the helix-turn-helix motif of AhpD, a signature motif found in the family of AhpD-like oxidoreductases [21][22][23] . The relative position of the motif within the primary sequence is similar between hSesn2 and YP_296737.1 but different in AhpD ( Supplementary Fig.…”

“…reductase 38,39 . However, the crystal structure of hSesn2 revealed that both Sesn-A and Sesn-C domains of hSesn2 bear the structural resemblance with R. eutropha YP_296737.1, which belongs to a family of alkylhydroperoxidases including M. tuberculosis AhpD 21 . Sequence alignment suggested that Sesn-A and YP_296737.1, but not Sesn-C, possess a limited sequence similarity with the catalytic active site sequence of M. tuberculosis AhpD (Supplementary Fig.…”

“…5a). The catalytic cysteine (Cys125) and residues in the proton delay system of AhpD-family oxidoreductases (Tyr127 and His132) are well-preserved within the Sesn-A domain 21 . Based on the structural similarity with bacterial alkylhydroxyperoxidases, we predicted that Sesn-A may function as a direct scavenger for ROS.…”

Transcriptome Analysis of the Intracellular Facultative Pathogen Piscirickettsia salmonis: Expression of Putative Groups of Genes Associated with Virulence and Iron Metabolism

“…The main strategy employed is the production of enzymes that degrade ROS species to maintain stress levels within a range of tolerance [91]. Our bacteria expressed alkyl hydroperoxidase D ( ahpD ) and alkyl hydroperoxide reductase C ( ahpC ), which provide significant antioxidant protection and have been described in various bacteria [92–94]. Additionally, Chung et al [95] demonstrated that deletion of the ahpC genes alone in V .…”