Biology:Diisopropyl-fluorophosphatase

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Diisopropyl-fluorophosphatase (EC 3.8.2.2, DFPase) refers to any enzyme able to catalyzes the reaction

  1. REDIRECT Template:Chemical reaction

This hydrolyses the nerve agent, diisopropyl fluorophosphate (DFP), to diisopropylphosphate and fluoride ion. Many examples are able to defeat a range of similar nerve agents used in chemical warfare.[1][2][3][4][5]

This enzyme is a hydrolase, specifically one acting on ester bonds phosphoric-triester hydrolases. The systematic name is diisopropyl-fluorophosphate fluorohydrolase. It requires a divalent cation such as calcium as a cofactor and is inhibited by chelating agents.[6] Other names in use include:[7]

  • Phrasing variants: diisopropyl­phosphofluoridase, dialkyll­fluorol­phosphatase, diisopropyl phosphorofluoridate hydrolase, isopropyll­phosphorol­fluoridase, and diisopropyll­fluorol­phosphonate dehalogenase.
  • Referring to other specific agents: tabunase, somanase.
  • More general terminology: organophosphorus acid anhydrolase (OPAA), organophosphate acid anhydrase, OPA anhydrase (see also EC 3.8.2.-, "Hydrolases; Acting on halide bonds; In P-halide compounds").

The kind of activity is found in a diverse range of organisms, including protozoa,[8] squid and clams,[9] mammals,[10] and soil bacteria.[11] Most examples of the enzyme also double as an aryldialkylphosphatase (EC 3.1.8.1), an enzyme that degrades arylated organophosphate compounds. DFPase was formally assigned EC 3.1.8.2 and aryldialkylphosphatase formally EC 3.8.2.1 – demonstrative of the close relationships of these enzymes.[12]

Because DFP (and other nerve agents) is not a natural compound, the ability to break it down is most likely an accident of evolution and not the native biological role of these enzymes. Some examples are apparantly involved in cellular dipeptide metabolism.[13][14]

Examples

The enzyme from Loligo vulgaris (European squid; Q7SIG4) head ganglion has been shown to detoxify a wide variety of similar nerve agents.[15][16]

A highly active form of the enzyme (pepQ; Q44238)is typically isolated from the marine bacteria Alteromonas undina for laboratory study. This form is both halophilic and thermophilic, making it particularly useful for detoxification applications.[17] Its main in vivo function seems to be that of a C-terminal Xaa-Pro dipeptidase (EC 3.4.13.9). It also has the ability to degrate non-halogenated organophosphates, activity.[18]

Less active variants of OPAA have also been isolated in slime mold, duckweed, and mung beans.[19]

Structure

Figure 2: pepQ trimer

The Alteromonas version, pepQ (also known as opaA), is a single polypeptide composed of 517 amino acids, with a molecular weight of 58 kDa.[20][14] The three-dimensional crystal structure of pepQ is a trimer, as seen in Figure 2. The enzyme has a 22% amino acid homology with human prolidase (PEPD) and a 30% homology to E. coli aminopeptidase P (P15034).[14] pepQ is unstable under harsh conditions, losing its activity in the presence of organic solvents, at elevated temperature, and over long-term storage.[1] Unprotected pepQ is also vulnerable to inhibition from other enzymes.[1] InterPro classifies it as a member of the Xaa-Pro dipeptidase family, IPR022846.[21]

The squid version has also been crystallized. Unlike pepQ it is a monomeric protein, a beta-propeller that comprises six blades: these two proteins have arrived at the same catalytic activity from very different origins.[22] InterPro classifies this enzyme as a member of the "SMP-30/Gluconolactonase/LRE-like region" family,[23] which includes human regucalcin and PON1, two other animal enzymes known to break down nerve agents.[24] Its endogenous function is unclear.

Structural studies

As of late 2007, 16 structures have been solved for this class of enzymes, with PDB accession codes 1E1A, 1PJX, 2GVU, 2GVV, 2GVW, 2GVX, 2IAO, 2IAP, 2IAQ, 2IAR, 2IAS, 2IAT, 2IAU, 2IAV, 2IAW, and 2IAX.

Mechanism

Figure 1: pepQ reaction mechanism

Although the exact mechanism of pepQ's nerve-agent detoxification is unclear, researchers have deduced that the basic reaction follows a general base mechanism with a simple in-line displacement of fluoride at the phosphorus center using an activated water molecule, as seen in Figure 1.[25]

pepQ activity is enhanced by reducing agents such as dithiothreitol (DTT) and beta-mercaptoethanol.[13] pepQ is also catalytically active over a wide pH range between 6.5 and 9.5 and temperature range between 10 and 65 °C, and is stimulated by manganese.[26]

Industrial relevance

OPAA’s susbstrate-specificity to organophosphorus-containing compounds, especially fluoride-containing G-type nerve agents such as sarin, cyclosarin, tabun, and soman,[27][28] have drawn the interest of the U.S. Army. That attention has resulted in the establishment of a biodegradation program in search of safe and effective means of disposal for chemical weapons in the 1980s; the program is currently called the Advanced Catalytic Enzyme System (ACES).[29]


References

  1. ↑ 1.0 1.1 1.2 Hoskin, F. C. G.; Rousch, A. H. (1982). "Hydrolysis of nerve gas by squid type diisopropylphosphorofluoridate hydrolyzing enzyme on agarose resin". Science 215 (4537): 1255–1257. doi:10.1126/science.7058344. PMID 7058344. Bibcode: 1982Sci...215.1255H. 
  2. ↑ Augustinsson K-B; Heimburger G (1954). "Enzymatic hydrolysis of organophosphorus compounds. II. Analysis of reaction products in experiments with Tabun and some properties of blood plasma tabunase". Acta Chem. Scand. 8: 762–767. doi:10.3891/acta.chem.scand.08-0762. 
  3. ↑ Augustinsson K-B; Heimburger G (1954). "Enzymatic hydrolysis of organophosphorus compounds. I. Occurrence of enzymes hydrolysing dimethyl-amido-ethoxy-phosphoryl cyanide (Tabun)". Acta Chem. Scand. 8: 753–761. doi:10.3891/acta.chem.scand.08-0753. 
  4. ↑ Augustinsson K-B; Heimburger G (1954). "Enzymatic hydrolysis of organophosphorus compounds. IV. Specificity studies". Acta Chem. Scand. 8: 1533–1541. doi:10.3891/acta.chem.scand.08-1533. 
  5. ↑ "Purification and properties of dialkylfluorophosphatase". Biochim. Biophys. Acta 26 (1): 29–39. 1957. doi:10.1016/0006-3002(57)90050-1. PMID 13479457. 
  6. ↑ Blum, Marc-Michael; Chen, Julian C.-H. (2010). "Structural characterization of the catalytic calcium-binding site in diisopropyl fluorophosphatase (DFPase)—Comparison with related β-propeller enzymes". Chemico-Biological Interactions 187 (1–3): 373–379. doi:10.1016/j.cbi.2010.02.043. PMID 20206152. 
  7. ↑ Enzyme 3.8.2.2 at KEGG Pathway Database.
  8. ↑ Landis, W. G.; Durst, H. D.; Savage Jr., R. E.; Haley, D. M.; Haley, M. V.; Johnson, D. W. (1987). "Discovery of multiple organofluorophosphate hydrolyzing activities in the protozoan Tetrahymena thermophila". J. Appl. Toxicol. 7 (1): 35–41. doi:10.1002/jat.2550070107. PMID 3611595. 
  9. ↑ Anderson, R. S.; Durst, H. D.; Landis, W. G. (1988). "Initial characterization of the organophosphate acid anhydrase activity in the clam, Rangia cuneata". Comp. Biochem. Physiol. 91C: 575–578. 
  10. ↑ Little, J. S., C. A. Broomfield, L. J. Boucher, and M. K. Fox-Talbot. 1986. Partial characterization of a rat liver enzyme that hydrolyzes sarin, soman, tabun and DFP. Fed. Proc. 45:791.
  11. ↑ DeFrank, J. J.; Cheng, T.-C. (1991). "Purification and properties of an organophosphorus acid anhydrase from a halophilic bacterial isolate". J. Bacteriol. 173 (6): 1938–1943. doi:10.1128/jb.173.6.1938-1943.1991. PMID 2001997. Bibcode: 1991JBact.173.1938D. 
  12. ↑ "ENZYME entry: EC 3.8.2.2". https://enzyme.expasy.org/EC/3.8.2.2. 
  13. ↑ 13.0 13.1 Elashvili, I, DeFrank, JJ, and Culotta, VC, Purification and characterization of DFPase from Alteromonas haloplanktis ATCC 14393, Proceedings of the 1998 ERDEC Scientific Conference on Chemical and Biological Defense Research, 17–20 November 1998, AD-A375 171.
  14. ↑ 14.0 14.1 14.2 Cheng, T-c, Liu, L, DeFrank, JJ, Anderson, D, and Hamilton, AB, Bacterial Nerve Agent Degrading Enzymes that are Similar to Prolidase Protein Sequence and Activity, Proceedings of the 1995 ERDEC Scientific Conference on Chemical Defense Research, 14–17 November 1995, AD-A315 812.
  15. ↑ Melzer, Marco; Chen, Julian C.-H.; Heidenreich, Anne et al. (2009). "Reversed Enantioselectivity of Diisopropyl Fluorophosphatase against Organophosphorus Nerve Agents by Rational Design". Journal of the American Chemical Society 131 (47): 17226–17232. doi:10.1021/ja905444g. PMID 19894712. 
  16. ↑ Li, Hexi; Lu, Cong; Liu, Zhenmin et al. (2024). "Advancements in bioscavenger mediated detoxification of organophosphorus poisoning". Toxicology Research 13 (3). doi:10.1093/toxres/tfae089. PMID 38863796. 
  17. ↑ Cheng, T.; Harvey, S.; Stroup, A. (1993). "Purification and properties of a highly active organophosphorus acid anhydrolase from alteromonas undina". Applied and Environmental Microbiology 59 (9): 3138–3140. doi:10.1128/AEM.59.9.3138-3140.1993. PMID 16349054. Bibcode: 1993ApEnM..59.3138C. 
  18. ↑ "UniProt - Q44238" (in en). https://www.uniprot.org/uniprotkb/Q44238/entry. 
  19. ↑ Hoskin, F.; Walker, J.; Mello, C. (1999). "Organophosphorus acid anhydrolase in slime mold, duckweed and mung bean: a continuing search for a physiological role and a natural substrate". Chemico-Biological Interactions 119–120: 399–404. doi:10.1016/S0009-2797(99)00051-4. PMID 10421476. Bibcode: 1999CBI...119..399H. 
  20. ↑ Cheng, T-c, Harvey, SP, and Chen, GL, Cloning and Expression of a Gene Encoding a Bacterial Enzyme for Decontamination of Organophosphorus Nerve Agents and Nucleotide Sequence of the Enzyme, Applied and Environmental Microbiology, 62 (1996) 1636-1641.
  21. ↑ "Q44238 Xaa-Pro dipeptidase". https://www.ebi.ac.uk/interpro/protein/UniProt/Q44238/. 
  22. ↑ Scharff, E. I.; Koepke, J.; Fritzsch, G.; Lücke, C.; Rüterjans, H. (June 2001). "Crystal structure of diisopropylfluorophosphatase from Loligo vulgaris". Structure 9 (6): 493–502. doi:10.1016/s0969-2126(01)00610-4. PMID 11435114. 
  23. ↑ "Q7SIG4 Diisopropyl-fluorophosphatase". https://www.ebi.ac.uk/interpro/protein/UniProt/Q7SIG4/. 
  24. ↑ "IPR013658 SMP-30/Gluconolactonase/LRE-like region". https://www.ebi.ac.uk/interpro/entry/InterPro/IPR013658/. 
  25. ↑ Dumas, DP; Caldwell, SR; Wild, JR; Raushel, FM (1989). "Purification and Properties of the Phosphotriesterase from Pseudomonas diminuta". Journal of Biological Chemistry 264 (33): 19659–19665. doi:10.1016/S0021-9258(19)47164-0. PMID 2555328. 
  26. ↑ Lewis, VE; Donarski, WJ; Wild, JR; Raushel, FM (1988). "Mechanism and Stereochemical course at phosphorus of the reaction catalyzed by a bacterial phosphotriesterase". Biochemistry 27 (5): 1591–1597. doi:10.1021/bi00405a030. PMID 2835095. 
  27. ↑ Cheng, T-c and Calomiris, JJ Stability of a nerve-agent-degrading enzyme in blast containment and firefighting foams, Proceedings of the 1994 ERDEC Scientific Conference on Chemical Defense Research, 15–18 November 1994, ADE 479952-001
  28. ↑ Cheng, T-c, DeFrank, JJ, Rastogi, VK, Alteromonas prolidase for the organophosphorus G-agent decontamination, Chemico-Biological Interactions 119-120(1999) 455-462.
  29. ↑ DeFrank, JJ, Cheng, T-c, Harvey, SP, and Rastogi, VK, Advanced catalytic enzyme system (ACES)-dual use capabilities, Proceedings of the 2002 Joint Service Scientific Conference on Chemical & Biological Defense Research, 19–21 November 2002, AD-M001 523.



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