Purification of a DFP-hydrolyzing enzyme from squid head ganglion

Hoskin, F. C. G.; Long, Ronald J.

doi:10.1016/0003-9861(72)90073-2

Cited by 50 publications

(16 citation statements)

References 24 publications

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“…Cleavage of the P-F or P-C bond of those compounds renders them relatively nontoxic as the bond broken is the one responsible for the irreversible inhibition of AChE (14). In contrast to this artificial function a physiological role of DFPase is not known yet but it can be excluded that squid-type DFPases function as ATPases or phosphatases (15). Squid-type DFPases are present in the optic ganglia, giant nerve axon, hepatopancreas, and salivary glants of cephalopods (16 -18).…”

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confidence: 87%

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High-Yield Expression, Purification, and Characterization of the Recombinant Diisopropylfluorophosphatase from Loligo vulgaris

Hartleib

Rüterjans

2001

Protein Expression and Purification

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confidence: 87%

“…DFPase activity was measured titrimetrically with a Radiometer "pH-Stat" apparature (15). The reaction was carried out in a volume of 3 ml at pH 7.5 and 25°C under a nitrogen atmosphere.…”

Section: Expression Of Protein Fragments Isolation Of Inclusion Bodimentioning

confidence: 99%

High-Yield Expression, Purification, and Characterization of the Recombinant Diisopropylfluorophosphatase from Loligo vulgaris

Hartleib

Rüterjans

2001

Protein Expression and Purification

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“…The mammalian serum paraoxonase (PON1) and squid diisopropylfluorophosphatase (DFPase) make up the final class of enzymes known to function as phosphotriesterases [11,12]. The X- ray structure of DFPase was solved to 0.85 Å with the wild-type enzyme, but the structure of PON1 could only be solved using an evolved variant with kinetic constants similar to the wild-type enzyme [20,78,79].…”

Section: β-Propeller Foldmentioning

confidence: 99%

Catalytic mechanisms for phosphotriesterases

Bigley

Raushel

2013

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

215

221

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Phosphotriesters are one class of highly toxic synthetic compounds known as organophosphates. Wide spread usage of organophosphates as insecticides as well as nerve agents has lead to numerous efforts to identify enzymes capable of detoxifying them. A wide array of enzymes has been found to have phosphotriesterase activity including phosphotriesterase (PTE), methyl parathion hydrolase (MPH), organophosphorus acid anhydrolase (OPAA), diisopropylfluorophosphatase (DFP), and paraoxonase 1 (PON1). These enzymes differ widely in protein sequence and three-dimensional structure, as well as in catalytic mechanism, but they also share several common features. All of the enzymes identified as phosphotriesterases are metal-dependent hydrolases that contain a hydrophobic active site with three discrete binding pockets to accommodate the substrate ester groups. Activation of the substrate phosphorus center is achieved by a direct interaction between the phosphoryl oxygen and a divalent metal in the active site. The mechanistic details of the hydrolytic reaction differ among the various enzymes with both direct attack of a hydroxide as well as covalent catalysis being found.

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“…In the past decades, a number of approaches have been investigated for protecting 1 3 138 Page 2 of 11 against OP compounds. A number of enzymes act as catalytic bioscavengers are considered to be an important potential strategy; especially, diisopropyl-fluorophosphatase (DFPase, EC 3.1.8.2, 35 kDa) from the head ganglion of squid Loligo vulgaris has received considerable attention [5][6][7]. Squid DFPase is shown to effectively catalyze the hydrolysis of the bond between phosphorus and the fluoride (or cyanide) leaving group (Scheme 1).…”

Section: Introductionmentioning

confidence: 99%

What roles do the residue Asp229 and the coordination variation of calcium play of the reaction mechanism of the diisopropyl-fluorophosphatase? A DFT investigation

Yang

et al. 2016

Theor Chem Acc

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much easier to perform the nucleophile attack. From the phosphoenzyme intermediate with the hexa-coordinated Ca 2+ , the uncoordinated Glu21 functions as a general base activated an additional water molecule to attack the carbon center of Asp229 and make the phosphate release. Residues Asn120 and Asn175 promote the elimination of the fluoride via donating strong hydrogen bonds. Residue Asp229 plays a dual role during the hydrolysis reaction process, either as a nucleophile or as a general base to activate the water nucleophile. The role of the calcium ion is providing a necessary conformation of the active site, facilitating the nucleophile formation and substrate orientation. Our calculated results shed further light on the organophosphate hydrolysis mechanism and guiding for structure-based protein engineering to modify hydrolysis rates and substrate specific.

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Purification of a DFP-hydrolyzing enzyme from squid head ganglion

Cited by 50 publications

References 24 publications

High-Yield Expression, Purification, and Characterization of the Recombinant Diisopropylfluorophosphatase from Loligo vulgaris

High-Yield Expression, Purification, and Characterization of the Recombinant Diisopropylfluorophosphatase from Loligo vulgaris

Catalytic mechanisms for phosphotriesterases

What roles do the residue Asp229 and the coordination variation of calcium play of the reaction mechanism of the diisopropyl-fluorophosphatase? A DFT investigation

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