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.      

FGF9 promotes survival of germ cells in the fetal testis

In addition to its role in somatic cell development in the testis, our data have revealed a role for Fgf9 in XY germ cell survival. In Fgf9-null mice, germ cells in the XY gonad decline in numbers after 11.5 days post coitum (dpc), while germ cell numbers in XX gonads are unaffected. We present evidence that germ cells resident in the XY gonad become dependent on FGF9 signaling between 10.5 dpc and 11.5 dpc, and that FGF9 directly promotes XY gonocyte survival after 11.5 dpc, independently from Sertoli cell differentiation. Furthermore, XY Fgf9-null gonads undergo true male-to-female sex reversal as they initiate but fail to maintain the male pathway and subsequently express markers of ovarian differentiation (Fst and Bmp2). By 14.5 dpc, these gonads contain germ cells that enter meiosis synchronously with ovarian gonocytes. FGF9 is necessary for 11.5 dpc XY gonocyte survival and is the earliest reported factor with a sex-specific role in regulating germ cell survival.     

QT-RR relationship in healthy subjects exhibits substantial intersubject variability and high intrasubject stability

Recently, it was demonstrated that the QT-RR relationship pattern varies significantly among healthy individuals. We compared the intra- and interindividual variations of the QT-RR relationship. Twenty-four-hour 12-lead digital electrocardiograms (ECGs; SEER MC, GE Marquette; 10-s ECG recorded every 30 s) were obtained at baseline and after 24 h, 1 wk, and 1 mo in 75 healthy subjects (42 women, 33 men, age 27.9 +/- 9.6 vs. 26.8 +/- 7.5 yr, P = not significant). QT interval was measured automatically in each ECG by six different algorithms, and the mean of the six measurements was analyzed. In each recording of each individual, QT-RR relationship was assessed by 10 different regression models including linear (QT = beta + alpha x RR) and parabolic (QT = beta x RR(alpha)) models. Standard deviations (SDs) of regression parameters alpha and beta of consecutive recordings of each individual were compared with SD of the individual means. Intrasubject stability and interindividual variability were further tested by ANOVA. With all models, intraindividual SDs of the regression parameters were highly significantly smaller than SD of individual means (P < 10(-5)-10(-9)). The intrasubject stability was further confirmed by ANOVA (P < 10(-19)-10(-30)). The QT-RR relationship exhibits substantial intersubject variability as well as a high intrasubject stability. This has practical implications for a precise estimation of the heart rate-corrected QT interval in which optimized subject-specific rate correction formulas should be used.     

Prolongation of the QT interval and post-extrasystolic augmentation of the TU-wave during emotional stress

We present a case of a 25-year-old woman with multiple blackouts and no structural heart disease, with abnormal T-U waves and borderline QT interval on her resting electrocardiogram. During emotional stress she developed frequent monomorphic ventricular premature beats, with characteristic changes of the sinus complexes immediately following the premature beats, namely augmentation and greater degree of merging of the T and U waves and QTc interval prolongation. The changes alert about the possibility of congenital long QT syndrome, specifically genotype 2 or 1.     

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.     

Mouse germ cell clusters form by aggregation as well as clonal divisions

After their arrival in the fetal gonad, mammalian germ cells express E-cadherin and are found in large clusters, similar to germ cell cysts in Drosophila. In Drosophila, germ cells in cysts are connected by ring canals. Several molecular components of intercellular bridges in mammalian cells have been identified, including TEX14, a protein required for the stabilization of intercellular bridges, and several associated proteins that are components of the cytokinesis complex. This has led to the hypothesis that germ cell clusters in the mammalian gonad arise through incomplete cell divisions. We tested this hypothesis by generating chimeras between GFP-positive and GFP-negative mice. We show that germ cell clusters in the fetal gonad arise through aggregation as well as cell division. Intercellular bridges, however, are likely restricted to cells of the same genotype.