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

<p>Abstract</p> <p>Background</p> <p>Alkyl 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.</p> <p>Results</p> <p>The crystal structure of a putative AhpD from <it>Pseudomonas aeruginosa </it>has been determined at 1.9 Å. The protein has an all-helical fold with a chain topology similar to a known AhpD from <it>Mycobacterium tuberculosis </it>despite a low overall sequence identity of 9%. A conserved two α-helical motif responsible for function is present in both. However, in the <it>P. aeruginosa </it>protein, helices H3, H4 of this motif are located at the N-terminal part of the chain, while in <it>M. tuberculosis </it>AhpD, the corresponding helices H8, H9 are situated at the C-terminus. Residues 24-62 of the putative catalytic region of <it>P. aeruginosa </it>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 <it>P. aeruginosa </it>exhibits a weak ability to reduce H₂O₂as tested using a ferrous oxidation-xylenol orange (FOX) assay, and this activity is blocked by thiol alkylating reagents.</p> <p>Conclusion</p> <p>Thus, 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.</p

Protein Engineering vol.9 no.9 pp.745-754, 19% Alternating charge clusters of side chains: new surface structural invariants observed in calf eye lens gamma-crystallins

A detailed stereochemical analysis of the oppositely charged side chains of amino acid residues on the surface of calf eye lens protein gamma-crystallin B has been carried out. The refined structural data of very high quality obtained at 1.47 A resolution have been used. Charge—charge interactions were considered to be valuable for all the charged oxygen and nitrogen atoms situated at distances, d, between 2.4 and 7.0 A. This means we consider short contact ion pairs as those with interchange distances 2.4 &lt; d ^s 4.0 A and distant contact ion pairs as those with distances 4.0 &lt; d = £ 7.0 A. JHydrogen bonding of the charged atomic groups with the structural water molecules also has been considered. We have not looked at the side groups of histidines which are charged only partially at neutral pH. Five clusters of charged side chains which were large enough were observed. The clusters are comprised of four to six residues which compose 543 % of the total charged residues in the protein. The clusters contain from eight to 12 charged atoms and look like the bent chains of oppositely charged atoms. All clusters are of plane geometry and their maximal linear dimensions are from 11 to 18 A. The root mean square deviations of charged atoms from the cluster plane varied from 0.63 to 0.86 A for four clusters and was only 1.85 A for the largest cluster. All clusters include a number of water molecules situated on the cluster boundary and grouped near the cluster plane. It was shown that the amino acid sequence positions of charged residues are conservative for all the proteins of the gamma-crystallin family of vertebrates including fish, frog, mouse, rat, calf and human. The cluster properties were discussed both in their functional aspect for gamma-crystallins and in other aspects common for globular proteins. As a result, the alternating charge clusters should be considered as newly recognized surface structural invariants. The importance of the charged side chain clusters is claimed for the updated concept of the protein surface

Novel approach for structural identification of protein family: glyoxalase I

Glyoxalase is one of two enzymes of the glyoxalase detoxification system against methylglyoxal and other aldehydes, the metabolites derived from glycolysis. The glyoxalase system is found almost in all living organisms: bacteria, protozoa, plants, and animals, including humans, and is related to the class of ‘life essential proteins’. The enzyme belongs to the expanded Glyoxalase/Bleomycin resistance protein/Dioxygenase superfamily. At present the GenBank contains about 700 of amino acid sequences of this enzyme type, and the Protein Data Bank includes dozens of spatial structures. We have offered a novel approach for structural identification of glyoxalase I protein family, which is based on the selecting of basic representative proteins with known structures. On this basis, six new subfamilies of these enzymes have been derived. Most populated subfamilies A1 and A2 were based on representative human Homo sapiens and bacterial Escherichia coli enzymes. We have found that the principle feature, which defines the subfamilies’ structural differences, is conditioned by arrangement of N- and C-domains inside the protein monomer. Finely, we have deduced the structural classification for the glyoxalase I and assigned about 460 protein sequences distributed among six new subfamilies. Structural similarities and specific differences of all the subfamilies have been presented. This approach can be used for structural identification of thousands of the so-called hypothetical proteins with the known PDB structures allowing to identify many of already existing atomic coordinate entrees

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

Abstract Background Alkyl 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. Results The 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. Conclusion Thus, 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