Glyceryl-Ether Monooxygenase [EC 1.14.16.5J Part VIII. Probing the Nature of the Active Site

Abstract

The V /K kinetic parameters for glyceryl-ether monooxygenase with 3-1 '-n-alkoxypropane-I,2-diols having alkyl sidechains varying from Cll to C22 (lb-k, 2a and b, and 3d) were determined. Maximum activity was found with the CI6 ethers, dropping almost to zero with ethers that have sidechains shorter than Cl2 and longer than C19. The thioether analogues with C16 and CI8 sidechains were poorer substrates than the respective oxygen ethers by one order of magnitude. The kinetic parameters for 2-1 '-n-hexadecyloxy-(6a) and 2-1 '-n-octadecyloxy-(6d) ethanol revealed that they were also better substrates than the respective thio-ethers by one order of magnitude and with these compounds those with C16 sidechains were better substrates than the C 18 analogues. L2-bis-3-1 '-n-Hexadecyloxypropan-3-01 (3c) and 1,3-bis-3-1'-n-hexadecyloxypropan-3-01 and 2-01 (4a) were inactive but 3-1'-n-hexadecyloxy-2-methoxypropan-I-ol (3b) and 3-1'-n-hexadecyloxy-l-methoxypropan-2-01 (4b) which have one long fatty chain were viable substrates because they possessed a free hydroxy group, but the ethers (6b, c and e), (7c and d) and (5) which do not have a free hydroxy group were inactive. These data, taken with those reported previously, have allowed us to postulate a model for the active site of the alkyl-ether substrates of the monooxygenase and to define the contours of the hydrophobic and hydrophilic pockets.Keywords: Kinetic parameters of Cll to C22 glycerol ethers, Cl6 and CI8 glycerol thioethers. ethylene glycol ethers and thioethers. syntheses. IH NMR, ORD. glyceryl-ether monooxygenase active site modeL alkyl-ether substrate domain.

Glyceryl-ether monooxygenase [EC 1.14.16.5]. A microsomal enzyme of ether lipid metabolism

“…The absolute configurations of the above three naturally occurring alcohols 2 are also consistent with optical rotatory dispersion studies. 30,31 On the rational assumption that glyceryl ethers, with a large variety of alkyl chains, from marine animals, 32,33 mammals and birds, 34 plants, 35 protozoa, 36 and bacteria 37,38 are formed by common biosynthetic pathways it is likely that they all have the same stereochemical configurations at C2 of the glycerol moiety. Early accounts on the history of glyceryl ether lipids have been admirably reviewed.…”

“…Today racemic isopropylidene glycerol, as well as its pure chiral enantiomers, are readily available commercially and have increased the popularity of this method considerably. Its metal salts (10) were prepared by reaction with sodium naphthalene, 13 dispersed sodium metal, 134 potassium metal, [135][136][137] and more recently sodium hydride (from a 60% dispersion in mineral oil) 31,138 in anhydrous inert solvents such as benzene, xylene, tetrahydrofuran or dimethyl formamide under dry nitrogen. After complete conversion to the salt (10), the reactive alkyl derivative such as the bromide, 138 137 in an inert solvent as above was added, and the mixture refluxed for several hours (4-48 h).…”

“…4%) by boiling for one hour or by setting aside overnight. 31 Enantiomerically pure glyceryl ethers (12, chiral at C2) were prepared from the corresponding chiral 1,2-isopropylidene glycerols, 13,26,31,139 and by using radiolabeled ( 14 C and/or 3 H) reactive alkyl derivatives and/or radioactive isopropylidene glycerol appropriately labeled, radiolabeled ␣-alkyl glycerols have been prepared. 136 No inversion of configuration occurs at the prochiral carbon atom C3 in (10) or (11) in this reaction, but inversion at the ␣-carbon atom of the reactive alkyl derivative does take place.…”

“…␣-Alkyl thioglyceryl ethers, on the other hand, were obtained in very high yields by boiling unprotected thioglycerol with alkyl bromides or iodides in the presence of potassium hydroxide in 95% ethanol (or methanol with a crystal of iodine), 138 or with the alkyl tosylate in the presence of triethylamine in ethanol. 31 In a modification of this procedure 35 S and/or 14 C labeled 3-hexadecylthio and 3-octadecylthio propane-1,2-diols were prepared from 35 S-thioglycerol or thioglycerol with the respective alkyl bromide or [1-14 C]-alkyl bromide dissolved in hexane in the presence of N-alcoholic potassium or sodium hydroxides under nitrogen. 145 A useful synthesis of chiral 1-O-alkyl-2-O-benzyl glycerols (which can be debenzylated by hydrogenolysis) starting from D-or L-1,2-isopropylidene-3-O-benzylthreitol was developed by Ohno et al 146 for the preparation of the enantiomers of PAF and related lipids.…”

“…Further alkylation of the 2-OH group (NaH/RX) provided 2,3-dialkoxy-1-benzyloxypropane from which 2,3-dialk-oxypropan-1-ol can be obtained by hydrogenolysis of the benzyl group. This propanol can also be further alkylated to furnish the racemic 1,2,3-trialkoxypropane, 31 and thus a variety of alkoxy groups can be introduced. If a variety of chiral dialkoxypropan-1-ols and trialkoxypropanes are required the methods developed by Eibl and coworkers 139 are very useful.…”