The Role of Interleukin-17, Interleukin-23, and Transforming Growth Factor-ß in Pregnancy Complicated by Placental Insufficiency.

AIM: The aim of the study was to evaluate the role of Interleukin-17 (IL-17), Interleukin-23 (IL-23), and transforming growth factor-ß (TGF-ß) in pregnancy complicated by placental insufficiency and in normal pregnancy. MATERIAL AND METHODS: The study comprised 34 patients with pregnancy complicated by fetal growth restriction (FGR) associated with preeclampsia (PE), as well as 35 healthy pregnant women. The concentrations of IL-17, IL-23, and TGF-ß in sera from maternal peripheral blood were determined by an immunoenzymatic assay. RESULTS: There were higher concentrations of IL-17 in the study group when compared to the controls. In the group of patients with placental insufficiency, the levels of IL-17 positively correlated with systolic blood pressure (R = 0.42, p < 0.01). The study obtained comparable concentrations of IL-23 in both studied groups. The concentrations of TGF-ß were significantly lower in pregnancy complicated by the insufficiency of placenta when compared to the controls. CONCLUSIONS: It seems possible that the increased concentrations of IL-17 and the deficiency of TGF-ß in pregnancy complicated by FGR and PE can be responsible for the activation of inflammatory response observed in PE cases.

The radical mechanism of biological methane synthesis by methyl-coenzyme M reductase.

Methyl-coenzyme M reductase, the rate-limiting enzyme in methanogenesis and anaerobic methane oxidation, is responsible for the biological production of more than 1 billion tons of methane per year. The mechanism of methane synthesis is thought to involve either methyl-nickel(III) or methyl radical/Ni(II)-thiolate intermediates. We employed transient kinetic, spectroscopic, and computational approaches to study the reaction between the active Ni(I) enzyme and substrates. Consistent with the methyl radical-based mechanism, there was no evidence for a methyl-Ni(III) species; furthermore, magnetic circular dichroism spectroscopy identified the Ni(II)-thiolate intermediate. Temperature-dependent transient kinetics also closely matched density functional theory predictions of the methyl radical mechanism. Identifying the key intermediate in methanogenesis provides fundamental insights to develop better catalysts for producing and activating an important fuel and potent greenhouse gas.

Crystal structures of S-HPCDH reveal determinants of stereospecificity for R- and S-hydroxypropyl-coenzyme M dehydrogenases.

(R)- and (S)-hydroxypropyl-coenzyme M dehydrogenases (R- and S-HPCDH) are stereospecific enzymes that are central to the metabolism of propylene and epoxide in Xanthobacter autotrophicus. The bacterium produces R- and S-HPCDH simultaneously to facilitate transformation of R- and S-enantiomers of epoxypropane to a common achiral product 2-ketopropyl-CoM (2-KPC). Both R- and S-HPCDH are highly specific for their respective substrates as each enzyme displays less than 0.5% activity with the opposite substrate isomer. In order to elucidate the structural basis for stereospecificity displayed by R- and S-HPCDH we have determined substrate bound crystal structures of S-HPCDH to 1.6Å resolution. Comparisons to the previously reported product-bound structure of R-HPCDH reveal that although the placement of catalytic residues within the active site of each enzyme is nearly identical, structural differences in the surrounding area provide each enzyme with a distinct substrate binding pocket. These structures demonstrate how chiral discrimination by R- and S-HPCDH results from alternative binding of the distal end of substrates within each substrate binding pocket.

Molecular basis for enantioselectivity in the (R)- and (S)-hydroxypropylthioethanesulfonate dehydrogenases, a unique pair of stereoselective short-chain dehydrogenases/reductases involved in aliphatic epoxide carboxylation.

(R)- and (S)-2-hydroxypropyl-CoM (R-HPC and S-HPC) are produced as intermediates in bacterial propylene metabolism from the nucleophilic addition of coenzyme M to (R)- and (S)-epoxypropane, respectively. Two highly enantioselective dehydrogenases (R-HPCDH and S-HPCDH) belonging to the short-chain dehydrogenase/reductase family catalyze the conversion of R-HPC and S-HPC to 2-ketopropyl-CoM (2-KPC), which undergoes reductive cleavage and carboxylation to produce acetoacetate. In the present study, one of three copies of S-HPCDH enzymes present on a linear megaplasmid in Xanthobacter autotrophicus strain Py2 has been cloned and overexpressed, allowing the first detailed side by side characterization of the R-HPCDH and S-HPCDH enzymes. The catalytic triad of S-HPCDH was found to consist of Y156, K160, and S143. R211 and K214 were identified as the amino acid residues coordinating the sulfonate of CoM in S-HPC. R211A and K214A mutants were severely impaired in the oxidation of S-HPC or reduction of 2-KPC but were largely unaffected in the oxidation and reduction of aliphatic alcohols and ketones. Kinetic analyses using R- and S-HPC as substrates revealed that enantioselectivity in R-HPCDH (value, 944) was dictated largely by differences in k(cat) while enantioselectivity for S-HPCDH (value, 1315) was dictated largely by changes in K(m). S-HPCDH had an inherent high enantioselectivity for producing (S)-2-butanol from 2-butanone that was unaffected by modulators that interact with the sulfonate binding site. The tertiary alcohol 2-methyl-2-hydroxypropyl-CoM (M-HPC) was a competitive inhibitor of R-HPCDH-catalyzed R-HPC oxidation, with a K(is) similar to the K(m) for R-HPC, but was not an inhibitor of S-HPCDH. The primary alcohol 2-hydroxyethyl-CoM was a substrate for both R-HPCDH and S-HPCDH with identical K(m) values. The pH dependence of kinetic parameters suggests that the hydroxyl group is a larger contributor to S-HPC binding to S-HPCDH than for R-HPC binding to R-HPCDH. It is proposed that active site constraints within the S-HPCDH prevent proper binding of R-HPC and M-HPC due to steric clashes with the improperly aligned methyl group on the C2 carbon, resulting in a different mechanism for controlling substrate specificity and enantioselectivity than present in the R-HPCDH.