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Fish Allergy
Published in Andreas L. Lopata, Food Allergy, 2017
Annette Kuehn, Karthik Arumugam
Enolases and aldolases are key enzymes of the catabolic glycolysis present in all tissues. Aldolase or 40 kDa-fructose-bisphosphate aldolase (EC 4.1.2.13) splits fructose 1,6-bisphosphate into triose phosphates dihydroxyacetone phosphate and glyceraldehyde 3-phosphate (4th step of glycolysis) (Garfinkel and Garfinkel 1985). Enolase or 50 kDa-phosphopyruvate hydratase (EC 4.2.1.11) is a metalloenzyme (Mg2+-ions per molecule) catalysing the conversion of 2-phosphoglycerate to phosphoenolpyruvate (9th step of glycolysis). Both enzymes belong to the structural family of so-called “TIM barrel”-proteins (Kuehn et al. 2016). Eponym for this family is the triosephosphate isomerase (TIM), which was characterized as the first protein by a common structure of eight alpha-helices alternating with eight beta-strands. Despite of the structural homology within this family, there is a lack of substantial sequence identity between TIM barrel-proteins.
Inorganic Chemical Pollutants
Published in William J. Rea, Kalpana D. Patel, Reversibility of Chronic Disease and Hypersensitivity, Volume 4, 2017
William J. Rea, Kalpana D. Patel
To understand the genetic and biochemical basis of microbial Hg methylation, Parks et al. analyzed the genomes of methylating and nonmethylating bacteria in the context of biochemical pathways involved in single-carbon metabolism. The well-characterized corrinoid iron–sulfur protein (CFeSP) is known to transfer methyl groups to a NiFeS cluster in acetyl-CoA synthase.571 Therefore, recognizing that a corrinoid protein associated with the acetyl-CoA pathway could be required for Hg methylation. They reasoned that a protein similar to CFeSP might transfer a methyl group to a Hg substrate to yield CH3Hg+, and that genes encoding such a protein should be recognizable in the genome sequences of Hg-methylating bacteria. Complete genome sequences are available for six methylating and eight closely related nonmethylating bacterial species (Tables 4.23 and 4.24). Furthermore, molecular structures and functions have been determined for various enzymes of the reductive acetyl-CoA pathway, including CFeSP from Moorella thermoacetica572 and Carboxdothermus hydrogenoformans.573,574 Accordingly, they performed a BLASTP search with the sequence of the large subunit of CFeSP (CfsA, locus tag CHY_1223 from C. hydrogenoformans Z-2901 against the translated genome sequence of D. desulfuricans ND 132).575 Sequence similarity was found between the C-terminal corrinoid-binding domain of DfsA and the N-terminus of DND132_1056, although DND132_1056 attacks both the TIM barrel domain and the C-terminal (4Fe-4S)-binding motif of CfsA. The C-terminal region showed no detectable similarity to any proteins of known structure, but exhibited features characteristic of a transmembrane domain.
Protein-Based Bioscavengers of Organophosphorus Nerve Agents
Published in Brian J. Lukey, James A. Romano, Salem Harry, Chemical Warfare Agents, 2019
Moshe Goldsmith, Yacov Ashani, Tamara. C. Otto, C. Linn Cadieux, David. S. Riddle
Phosphotriesterase (PTE), also termed organophosphate hydrolase (OPH) (EC 3.1.8.1), is a 336-amino acid (36 kDa), zinc-dependent hydrolase, structured as an (αβ)8 TIM-barrel (Benning et al., 1994). It is a bacterial enzyme that belongs to the amidohydrolase superfamily and was first identified in soil bacteria that hydrolyzed the pesticide parathion (Serdar et al., 1982). Since man-made OP pesticides were introduced into the environment only in the 1950s, and since its catalytic efficiency of hydrolysis approaches the diffusion limit (e.g., kcat/KM ~ 109 M−1 min−1 with paraoxon), it was suggested that PTE is the product of the recent and rapid natural evolution of a lactonase (Afriat-Jurnou et al., 2012; Afriat et al., 2006). PTE is encoded on a natural plasmid (pCMS1) in a bacterial strain that was originally classified as Pseudomonas diminuta strain MG (Serdar and Gibson, 1985; Serdar et al., 1982) and later reclassified as Brevundimonas diminuta (Segers et al., 1994). The fact that the gene encoding PTE (opd) was found on a plasmid meant that it could be transferred to other bacterial strains. Indeed, PTE was also found independently in unrelated soil bacteria isolated from different parts of the world, such as Sphingobium fuliginis (former Flavobacterium sp. ATCC 27551) identified in the Philippines (Kawahara et al., 2010; Mulbry and Karns, 1989; Sethunathan and Yoshida, 1973) and Brevundimonas diminuta and Pseudomonas putida, identified in the United States (Iyer et al., 2013b; Serdar et al., 1982). Plasmid pCMS1 from B. diminuta was found to be self-transmissible and responsible for the horizontal transfer of its opd gene between soil bacteria (Pandeeti et al., 2011). OP pesticides can become a valuable source of phosphorus for soil bacteria. However, they need to be imported into the cell and degraded to simpler molecules, such as phosphoric acid, to be used for growth and energy. Accordingly, the expression of PTE from B. diminuta was found to be targeted to the inner membrane of the bacteria, where it becomes membrane bound and associated with phosphatases and with a phosphate transporter (Parthasarathy et al., 2016).
Novel and potent inhibitors for dihydropteroate synthase of Helicobacter pylori
Published in Journal of Receptors and Signal Transduction, 2020
Sri Harsha Satuluri, Sudheer Kumar Katari, Chiranjeevi Pasala, Umamaheswari Amineni
DHPS structure is highly conserved and consists of two identical monomers of a classical triosephosphate isomerase (TIM) type barrel. This TIM barrel consists of eight parallel beta strands encircled by eight alpha helices. The active site of DHPS monomer is located at the C-terminal end of the β-barrel which comprises three conserved sub-site: the pterin-binding site, the pABA-binding site, and the anion-binding site. The pterin-binding pocket is situated in a deep cleft of the C-terminal end of β-barrel, whereas the pABA-binding pocket is located at the surface and comprises of two flexible loops (loop1 and loop2) [25,26]. Initially, DHPP binds with DHPS active site and it catalyzes the slow release of pyrophosphate moiety from DHPP [16]. The released pyrophosphate moiety plays an important role in stabilizing the conformation of flexible pABA-binding loops. After pABA-binding, the condensation of DHPP + and pABA occurs through SN1 reaction to form dihydropteroate. The mutation in loop 1 and loop 2 allow the binding of pABA but disrupt the binding of sulfa compounds [16].
Hydrogen deuterium exchange mass spectrometry applied to chaperones and chaperone-assisted protein folding
Published in Expert Review of Proteomics, 2019
Florian Georgescauld, Thomas E. Wales, John R. Engen
The folding of a natural GroEL/GroES substrate was analyzed by HDX MS for the TIM-barrel protein dihydrodipicolinate synthase (DAPA) [52]. DAPA is a tetrameric enzyme whose folding is strictly dependent on GroEL/GroES at 37°C. At a lower temperature, spontaneous folding of DAPA without chaperone assistance is possible, allowing a comparison of the folding pathway of spontaneous DAPA folding with that of GroEL/GroES-assisted folding (Figure 8). Protection from HDX as a result of monomer folding versus assembly of the final tetrameric form could be distinguished. During spontaneous folding, HDX MS showed that peptides in the TIM-barrel refolded with an identical slow t1/2 of 9 min, a rate that was also obtained in completely independent enzyme assays. These results indicated that the TIM-barrel domain acquires its native state in a single slow step. HDX MS showed the strong acceleration of folding by GroEL/GroES, especially evident in the TIM-barrel domain; a single 30 s encapsulation cycle inside GroEL/GroES was sufficient for renaturation although tetrameric assembly was slower and matched assembly values obtained by enzymatic assays. The folding pathway with GroEL was different than spontaneous folding in that several subdomains of the beta barrel acquired their native structure at different speeds (Figure 8). These HDX MS results demonstrated that the folding pathway of a protein is modified when it is encapsulated inside the GroEL/GroES chaperone, resulting in folding acceleration.
Advances in detection of hazardous organophosphorus compounds using organophosphorus hydrolase based biosensors
Published in Critical Reviews in Toxicology, 2019
Monika Jain, Priyanka Yadav, Abhijeet Joshi, Prashant Kodgire
PTE/OPH, a homodimer metalloprotein, is a member of superfamily amidohydrolase. It utilizes a divalent ion, such as Co2+, Zn2+, Mg2+, Ca2+, and Fe2+ etc., for the nucleophilic attack by activating the hydrolytic water molecules (Ghanem and Raushel 2005). With respect to the normal activity, enzymes supplemented with these ions show 121, 86, 81, 74, and 48% activity, respectively. The high-resolution X-ray structure showed that the protein folds into αβ barrel motif also known as TIM Barrel (Raushel 2002). OPH adopts TIM barrel fold where the active site is located at the C terminal of the β barrel (Ghanem and Raushel 2005). Two Zn2+ are bound to the histidine residues at the binuclear metallic center. One zinc cation is bound to two histidine residues, H201 from β strand 5 and H206 from β strand 6. Another zinc cation is bound to two histidine residues (H55 and H57) and one aspartic acid (D301) (Ghanem and Raushel 2005; Bigley and Raushel 2013). Carboxylated lysine from β strand 4 and hydroxide ion form the bridge between the two cations (Bigley and Raushel 2013).