Purification and characterization of 3-dehydroquinase from Escherichia coli.

A procedure has been developed for the purification of 3-dehydroquinase from Escherichia coli. Homogeneous enzyme with specific activity 163 units/mg of protein was obtained in 19% overall yield. The subunit Mr estimated from polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate was 29,000. The native Mr, estimated by gel permeation chromatography on Sephacryl S-200 (superfine) and on TSK G3000SW, was in the range 52,000-58,000, indicating that the enzyme is dimeric. The catalytic properties of the enzyme have been determined and shown to be very similar to those of the biosynthetic 3-dehydroquinase component of the arom multifunctional enzyme of Neurospora crassa.     

Triterpenoid saponins from the roots of tea plant (Camellia sinensis var. assamica)

Three olean-12-ene type triterpenoid saponins, named TR-saponins A, B and C, were isolated as methyl esters from tea roots (Camellia sinesis var. assamica) after treatment with diazomethane. Their structures were established as the methyl esters of 3-O-alpha-L-arabinopyranosyl (1-->3)-beta-D-glucuronopyranosyl-21, 22-di-O-angeloyl-R1-barrigenol-23-oic acid, 3-O-alpha-L-arabinopyranosyl (1-->3)-beta-D-glucuronopyranosyl-21-O-angeloyl-22-O-2-me thylbutanoyl-R1- barrigenol-23-oic acid and 3-O-alpha-L-arabinopyranosyl (1-->3)-beta-D-glucuronopyranosyl-16 alpha-O-acetyl-21-O-angeloyl-22-O-2-methylbutanoyl-R1-bar rigenol-23-oic acid, by extensive 1D and 2D-NMR as well as FABMS and HR-MS analyses.     

Thioredoxin 2 haploinsufficiency in mice results in impaired mitochondrial function and increased oxidative stress

The mitochondrial form of thioredoxin, thioredoxin 2 (Txn2), plays an important role in redox control and protection against ROS-induced mitochondrial damage. To evaluate the effect of reduced levels of Txn2 in vivo, we measured oxidative damage and mitochondrial function using mice heterozygous for the Txn2 gene (Txn2(+/-)). The Txn2(+/-) mice showed approximately 50% decrease in Trx-2 protein expression in all tissues without upregulating the other major components of the antioxidant defense system. Reduced levels of Txn2 resulted in decreased mitochondrial function as shown by reduced ATP production by isolated mitochondria and reduced activity of electron transport chain complexes (ETCs). Mitochondria isolated from Txn2(+/-) mice also showed increased ROS production compared to wild type mice. The Txn2(+/-) mice showed increased oxidative damage to nuclear DNA, lipids, and proteins in liver. In addition, we observed an increase in apoptosis in liver from Txn2(+/-) mice compared with wild type mice after diquat treatment. Our results suggest that Txn2 plays an important role in protecting the mitochondria against oxidative stress and in sensitizing the cells to ROS-induced apoptosis.     

Activation of brain phenylethanolamine-N-methyltransferase by Triton-X-100: Time related differences in the enzyme activation

The effect of different concentrations of Triton-X-100 (0.2 to 5 %) on the activity of enzyme phenylethanolamine-N-methyltransferase (PNMT) in the brain and adrenal was studied. The addition of 0.2 % Triton-X-100 to the 0.9 % KCl homogenization media resulted in 180 % activation of the brain PNMT. The similar content of this detergent added to the adrenal PNMT preparation had no marked effect on enzyme activity. Rising Triton-X-100 concentrations from 0.2 % to 5 % resulted in higher activation of brain PNMT activity but the adrenal enzyme remained rather stable. An exposure of 15 minutes of brain PNMT preparation to Triton-X-100 was the optimal interval to evoke the maximal increase in enzyme activity. This activation of brain PNMT by Triton-X-100 was observable up to 24 hours after the addition of the detergent.     

Mitochondrial haplogroup N1a phylogeography,with implication to the origin of European farmers

Background  

Tracing the genetic origin of central European farmer N1a lineages can provide a unique opportunity to assess the patterns of the farming technology spread into central Europe in the human prehistory. Here, we have chosen twelve N1a samples from modern populations which are most similar with the farmer N1a types and performed the complete mitochondrial DNA genome sequencing analysis. To assess the genetic and phylogeographic relationship, we performed a detailed survey of modern published N1a types from Eurasian and African populations.     

The Internal Organization of Mycobacterial Partition Assembly: Does the DNA Wrap a Protein Core?

Before cell division in many bacteria, the ParBs spread on a large segment of DNA encompassing the origin-proximal parS site(s) to form the partition assembly that participates in chromosome segregation. Little is known about the structural organization of chromosomal partition assembly. We report solution X-ray and neutron scattering data characterizing the size parameters and internal organization of a nucleoprotein assembly formed by the mycobacterial chromosomal ParB and a 120-meric DNA containing a parS-encompassing region from the mycobacterial genome. The cross-sectional radii of gyration and linear mass density describing the rod-like ParB-DNA assembly were determined from solution scattering. A “DNA outside, protein inside” mode of partition assembly organization consistent with the neutron scattering hydrogen/deuterium contrast variation data is discussed. In this organization, the high scattering DNA is positioned towards the outer region of the partition assembly. The new results presented here provide a basis for understanding how ParBs organize the parS-proximal chromosome, thus setting the stage for further interactions with the DNA condensins, the origin tethering factors and the ParA.     

Differentiation of Isomeric Purine and Pyrimidine Mononucleotides by Fast Atom Bombardment Tandem Mass Spectrometry

Abstract

The use of positive ion fast atom bombardment mass-analysed ion kinetic energy (FAB/MIKE) spectroscopy to differentiate the 2′, 3′-and 5′-monophosphate isomers of adenosine, guanosine and cytidine is described.