An essential saccharide binding domain for the mAb 2C7 established for Neisseria gonorrhoeae LOS by ES-MS and MSn

A study of bacterial surface oligosaccharides were investigated among different strains of Neisseria gonorrhoeae to correlate structural features essential for binding to the MAb 2C7. This epitope is widely expressed and conserved in gonococcal isolates, characteristics essential to an effective candidate vaccine antigen. Sample lipooligosaccharides (LOS), was prepared by a modification of the hot phenol-water method from which de-O-acetylated LOS and oligosaccharide (OS) components were analyzed by ES-MS-CID-MS and ES-MSnin a triple quadrupole and an ion trap mass spectrometer, respectively. Previously documented natural heterogeneity was apparent from both LOS and OS preparations which was admixed with fragments induced by hydrazine and mild acid treatment. Natural heterogeneity was limited to phosphorylation and antenni extensions to the alpha-chain. Mild acid hydrolysis to release OS also hydrolyzed the beta(1-->6) glycosidic linkage of lipid A. OS structures were determined by collisional and resonance excitation combined with MS and multistep MSn which provided sequence information from both neutral loss, and nonreducing terminal fragments. A comparison of OS structures, with earlier knowledge of MAb binding, enzyme treatment, and partial acid hydrolysis indicates a generic overlapping domain for 2C7 binding. Reoccurring structural features include a Hepalpha(1-->3)Hepbeta(1-->5)KDO trisaccharide core branched on the nonreducing terminus (Hep-2) with an alpha(1-->2) linked GlcNAc (gamma-chain), and an alpha-linked lactose (beta-chain) residue. From the central heptose (Hep-1), a beta(1-->4) linked lactose (alpha-chain), moiety is required although extensions to this residue appear unnecessary.      

The Role of Preventing Nitric Oxide Deficiency in the Antihypertensive Effect of Adaptation to Hypoxia

Shortage of endothelial nitric oxide (NO) manifested as decreased daily urinary excretion of nitrate and nitrite as well as attenuated endothelium-dependent relaxation of conduit and resistance vessels progresses with age-related increase of blood pressure (BP) in stroke-prone spontaneously hypertensive rats (SHRSP). Simultaneous NO-dependent suppression of vascular contractions is, apparently, due to the inducible NO synthase activity in vascular smooth muscle specific for spontaneously hypertensive rat. The adaptation of rats to hypobaric hypoxia initiated at early hypertensive stage (at the age of 5–6 weeks) decelerates hypertension progress. The antihypertensive effect of the adaptation was accompanied by stimulation of endothelial NO synthesis and prevention of impaired NO-dependent response in isolated blood vessels. Nitric oxide stores were formed in the vascular wall of SHRSP and WKY rats at the same time. The obtained data indicate that the correction of endothelial NO deficiency plays a significant role in the antihypertensive effect of adaptation to hypoxia.     

Role of nitric oxide in adaptation to hypoxia and adaptive defense

Adaptation to hypoxia is beneficial in cardiovascular pathology related to NO shortage or overproduction. However, the question about the influence of adaptation to hypoxia on NO metabolism has remained open. The present work was aimed at the relationship between processes of NO production and storage during adaptation to hypoxia and the possible protective significance of these processes. Rats were adapted to intermittent hypobaric hypoxia in an altitude chamber. NO production was determined by plasma nitrite/nitrate level. Vascular NO stores were evaluated by relaxation of the isolated aorta to diethyldithiocarbamate. Experimental myocardial infarction was used as a model of NO overproduction; stroke-prone spontaneously hypertensive rats (SHR-SP) were used as a model of NO shortage. During adaptation to hypoxia, the plasma nitrite/nitrate level progressively increased and was correlated with the increase in NO stores. Adaptation to hypoxia prevented the excessive endothelium-dependent relaxation and hypotension characteristic for myocardial infarction. At the same time, the adaptation attenuated the increase in blood pressure and prevented the impairment of endothelium-dependent relaxation in SHR-SP. The data suggest that NO stores induced by adaptation to hypoxia can either bind excessive NO to protect the organism against NO overproduction or provide a NO reserve to be used in NO deficiency.     

Correction of NO-dependent cardiovascular disorders by adaptation to hypoxia

Spontaneously hypertensive rats (SHR-SP) were adapted to intermittent hypobaric hypoxia in an altitude chamber for 40 days. The adaptation to hypoxia prevented an excessive endothelium-dependent relaxation and hypotension characteristic of myocardial infarction. The adaptation also attenuated the increase in blood pressure and prevented impairment of the endothelium-dependent relaxation in SHR-SP. The universal nature of the adaptation allows to use it for correcting many cardiovascular disorders related to diverse alterations of NO metabolism.     

Effect of self-association on the structural organization of partially folded proteins: inactivated actin

The propensity to associate or aggregate is one of the characteristic properties of many nonnative proteins. The aggregation of proteins is responsible for a number of human diseases and is a significant problem in biotechnology. Despite this, little is currently known about the effect of self-association on the structural properties and conformational stability of partially folded protein molecules. G-actin is shown to form equilibrium unfolding intermediate in the vicinity of 1.5 M guanidinium chloride (GdmCl). Refolding from the GdmCl unfolded state is terminated at the stage of formation of the same intermediate state. An analogous form, known as inactivated actin, can be obtained by heat treatment, or at moderate urea concentration, or by the release of Ca(2+). In all cases actin forms specific associates comprising partially folded protein molecules. The structural properties and conformational stability of inactivated actin were studied over a wide range of protein concentrations, and it was established that the process of self-association is rather specific. We have also shown that inactivated actin, being denatured, is characterized by a relatively rigid microenvironment of aromatic residues and exhibits a considerable limitation in the internal mobility of tryptophans. This means that specific self-association can play an important structure-forming role for the partially folded protein molecules.