, Volume 66, Issue 3, pp 285-290
Date: 09 Oct 2004

Carbon isotope fractionation during cistrans isomerization of unsaturated fatty acids in Pseudomonas putida

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Abstract

The molecular mechanism of the unique cis to trans isomerization of unsaturated fatty acids in the solvent-tolerant bacterium Pseudomonas putida S12 was studied. For this purpose, the carbon isotope fractionation of the cistrans isomerase was estimated. In resting cell experiments, addition of 3-nitrotoluene for activation of the cistrans isomerase resulted in the conversion of the cis-unsaturated fatty acids into the corresponding trans isomers. For the conversion of C16:1 cis to its corresponding trans isomer, a significant fractionation was measured. The intensity of this fractionation strongly depended on the rate of cistrans isomerization and the added concentration of 3-nitrotoluene, respectively. The presence of a significant fractionation provides additional indication for a transition from the sp2 carbon linkage of the cis-double bond to an intermediate sp3 within an enzyme–substrate complex. The sp2 linkage is reconstituted after rotation to the trans configuration has occurred. As cytochrome c plays a major role in the catabolism of Cti polypeptide, these findings favour a mechanism for the enzyme in which electrophilic iron (Fe3+), provided by a heme domain, removes an electron of the cis double bond thereby transferring the sp2 linkage into sp3.