The organophosphorus nerve agents sarin, soman, tabun, and VX exert their toxic effects by inhibiting the action of human acetylcholinesterase, a member of the serine hydrolase superfamily of enzymes. The current treatments for nerve agent exposure must be administered quickly to be effective and they often do not eliminate long-term toxic side effects associated with organophosphate poisoning. Thus, there is significant need for effective prophylactic methods to protect at-risk personnel from nerve agent exposure, and protein-based approaches have emerged as promising candidates. We present the 2.7 Å resolution crystal structures of the serine hydrolase human carboxylesterase 1 (hCE1), a broad-spectrum drug metabolism enzyme, in covalent acylenzyme intermediate complexes with the chemical weapons soman and tabun. The structures reveal that hCE1 binds stereoselectively to these nerve agents; for example, hCE1 appears to react preferentially with the 10 4 -fold more lethal P S stereoisomer of soman relative to the P R form. In addition, structural features of the hCE1 active site indicate that the enzyme may be resistant to deadend organophosphate aging reactions that permanently inactivate other serine hydrolases. Taken together, these data provide important structural details toward the goal of engineering hCE1 into an organophosphate hydrolase and protein-based therapeutic for nerve agent exposure.The organophosphorus (OP) nerve agents sarin, soman, tabun, and VX are among the deadliest man-made chemicals (1). While the military use of OP nerve agents is widely banned, these compounds have been employed in recent decades by rogue states and terrorist groups. In 1988, Iraq employed weaponized sarin against its own Kurdish citizens in Halabja, a town adjacent to the Iranian border, killing an estimated 5,000 (2). Coordinated attacks on the Tokyo subway system by the Japanese Aum Shinrikyo cult in 1995 also employed sarin, killing 12 and injuring † This work was supported, in part, by NIH grants CA98468 and NS58089, and the American Lebanese Syrian Associated Charities. ‡ Protein Data Bank codes are 2HRQ for the hCE1-soman structure and 2HRR for the hCE1-tabun structure. *Corresponding Author: Matthew R. Redinbo, Ph.D., Department of Chemistry, CB #3290, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-3290, (919)843-8910; Fax: (919)962-2388, redinbo@unc.edu. 1 Abbreviations: AcChE, Human acetylcholinesterase; BuChE, Human butyrylcholinesterase; CE, carboxylesterase; hCE1, Human carboxylesterase 1; HI-6, 1-(2-hydroxy-iminomethylpyridinium)-1-(4-carboxyamino)-pyridinium dimethylether dichloride; MuAcChE, Mus musculus acetylcholinesterase; OP, organophosphate; Ortho-7, 1,7-heptylene-bis-N,N′-2-pyridiniumaldoxime dichloride; PON1, human serum paraoxonase 1; rmsd, root mean square deviation; Sarin, methylethyl methylphosphonofluoridate; Soman, Tabun, ethyl N, TcAcChE, Torpedo californica acetylcholinesterase; VX, OP toxicity is thought to be mediated through the inhibition of human acety...

The Recovery Phase Following a Triple Iron TriathlonThe purpose of this case study was to investigate the recovery phase in a single athlete after a Triple Iron Triathlon involving 11.4 km swimming, 540 km cycling and 126.6 km running. Total body mass, body fat and skeletal muscle mass using the anthropometric method as well as total body water using bioelectrical impedance analysis were determined pre race, after the race and every 24 hours until complete recovery. Parameters of hydration status (urinary specific gravity, hematocrit and plasma sodium) and skeletal muscle damage (plasma urea) were measured at the same time. After finishing the race within 42 hours, total body mass was decreased and total body water was increased. Over the following 6 days, prior to returning to pre race values for plasma volume and total body water, body mass reached a peak value on day 3, plasma volume on day 2 and total body water on day 1. Clinically visible edemas of the feet persisted until day 4. Six days after the race, body mass was reduced by 2.1 kg, skeletal muscle mass by 0.6 kg and fat mass by 0.7 kg. An increase in both blood urea and urinary output post race between days 3 and 6 suggested an impairment of renal function immediately post race due to skeletal muscle damage and manifesting clinically observed edemas. For practical application, athletes, coaches and physicians should anticipate that performing such an ultra-endurance race can lead to considerable edemas of the lower limbs during the recovery phase.

The enzyme group-VIIA phospholipase A2 (gVIIA-PLA2) is bound to lipoproteins in human blood and hydrolyzes the ester bond at the sn-2 position of phospholipid substrates with a short sn-2 chain. The enzyme belongs to a serine hydrolase superfamily of enzymes, which react with organophosphorus (OP) nerve agents. OPs ultimately exert their toxicity by inhibiting human acetycholinesterase at nerve synapses, but may additionally have detrimental effects through inhibition of other serine hydrolases. We have solved the crystal structures of gVIIA-PLA2 following inhibition with the OPs diisopropylfluorophosphate, sarin, soman and tabun. The sarin and soman complexes displayed a racemic mix of P R and P S stereoisomers at the P-chiral center. The tabun complex displayed only the P R stereoisomer in the crystal. In all cases, the crystal structures contained intact OP adducts that had not aged. Aging refers to a secondary process OP complexes can go through, which dealkylates the nerve agent adduct and results in a form that is highly resistant to either spontaneous or oxime-mediated reactivation. Non-aged OP complexes of the enzyme were corroborated by trypsin digest and matrix assisted laser desorption ionization mass spectrometry of OP-enzyme complexes. The lack of stereoselectivity of sarin reaction was confirmed by gas chromatography/mass spectrometry using a chiral column to separate and quantitate the unbound stereoisomers of sarin following incubation with enzyme. The structural details and characterization of nascent reactivity of several toxic nerve agents is discussed with a long term goal of developing gVIIA-PLA2 as a catalytic bioscavenger of OP nerve agents.Keywords phospholipase A2; Lp-PLA2; PAF-AH; organophosphate; nerve agent ☆ Protein Data Bank entry codes 3F9C, 3F96, 3F97, and 3F98 are non-aged complexes of human group-VIIA phospholipase A2 with the pesticide diisopropylfluorophosphate or the nerve agents sarin, soman and tabun, respectively.