Quaternary organization of the goodpasture autoantigen,the alpha 3(IV) collagen chain. Sequestration of two cryptic autoepitopes by intrapromoter interactions with the alpha4 and alpha5 NC1 domains

Goodpasture's (GP) disease is caused by autoantibodies that target the alpha3(IV) collagen chain in the glomerular basement membrane (GBM). Goodpasture autoantibodies bind two conformational epitopes (E(A) and E(B)) located within the non-collagenous (NC1) domain of this chain, which are sequestered within the NC1 hexamer of the type IV collagen network containing the alpha3(IV), alpha4(IV), and alpha5(IV) chains. In this study, the quaternary organization of these chains and the molecular basis for the sequestration of the epitopes were investigated. This was accomplished by physicochemical and immunochemical characterization of the NC1 hexamers using chain-specific antibodies. The hexamers were found to have a molecular composition of (alpha3)(2)(alpha4)(2)(alpha5)(2) and to contain cross-linked alpha3-alpha5 heterodimers and alpha4-alpha4 homodimers. Together with association studies of individual NC1 domains, these findings indicate that the alpha3, alpha4, and alpha5 chains occur together in the same triple-helical protomer. In the GBM, this protomer dimerizes through NC1-NC1 domain interactions such that the alpha3, alpha4, and alpha5 chains of one protomer connect with the alpha5, alpha4, and alpha3 chains of the opposite protomer, respectively. The immunodominant Goodpasture autoepitope, located within the E(A) region, is sequestered within the alpha3alpha4alpha5 protomer near the triple-helical junction, at the interface between the alpha3NC1 and alpha5NC1 domains, whereas the E(B) epitope is sequestered at the interface between the alpha3NC1 and alpha4NC1 domains. The results also reveal the network distribution of the six chains of collagen IV in the renal glomerulus and provide a molecular explanation for the absence of the alpha3, alpha4, alpha5, and alpha6 chains in Alport syndrome.     

Blue, red and blue plus red light control of chlorophyll content and CO2 gas exchange in barley leaves: Quantitative description of the effects of light quality and fluence rate

The dependence of the Chl content and the rate of CO₂ gas exchange (RGE) on both blue and red quanta fluence rates have been studied in primary leaves of barley ( Hordeum vulgare L. cv. Viner). Empirical equations connecting the two photosynthetic indices with fluence rates of blue or red light were developed. These equations consist of 3 (Chl content) or 2 (RGE) terms, each reflecting the involvement of a specific reaction in the long-term light control of the development of the photosynthetic apparatus. On the basis of the equations the effects of mixed blue plus red light on both the Chl content and RGE were calculated. An additive mode of the co-action of blue and red light in the range of high PFDs (10–170 μmol m⁻² s⁻¹) becomes evident from the comparison of the experimental results and calculated data. The results indicate the involvement of phytochrome, cryptochrome and chlorophyll in the long-term regulation of the Chl content and RGE.     

19F NMR study on the biodegradation of fluorophenols by various Rhodococcus species

Of all NMR observable isotopes 19F is the one perhaps most convenient for studies on biodegradation of environmental pollutants. The reasons underlying this potential of 19F NMR are discussed and illustrated on the basis of a study on the biodegradation of fluorophenols by four Rhodococcus strains. The results indicate marked differences between the biodegradation pathways of fluorophenols among the various Rhodococcus species. This holds not only for the level and nature of the fluorinated biodegradation pathway intermediates that accumulate, but also for the regioselectivity of the initial hydroxylation step. Several of the Rhodococcus species contain a phenol hydroxylase that catalyses the oxidative defluorination of ortho-fluorinated di- and trifluorophenols. Furthermore, it is illustrated how the 19F NMR technique can be used as a tool in the process of identification of an accumulated unknown metabolite, in this case most likely 5-fluoromaleylacetate. Altogether, the 19F NMR technique proved valid to obtain detailed information on the microbial biodegradation pathways of fluorinated organics, but also to provide information on the specificity of enzymes generally considered unstable and, for this reason, not much studied so far.     

Recombinant human peroxiredoxin VI: preparation and protective properties in vitro

cDNA of human peroxiredoxin VI, one of the recently discovered novel antioxidant proteins, was expressed in Escherichia coli cells. The expression product was obtained in water-soluble form and purified by a two-step chromatographic procedure using DEAE-Sepharose and Sephacryl S-200. According to CD data, the polypeptide chain of the recombinant human peroxiredoxin VI contains 40% -helical region and 30% -structure, which is the same as for native rat peroxiredoxin VI. The protective properties of the recombinant protein determined as its ability to prevent the inactivation of glutamine synthetase from E. coli in a model oxidation system were comparable with the protective properties of native rat peroxiredoxin VI.     

Extracellular Oxidase from the Neonothopanus nambi Fungus as a Promising Enzyme for Analytical Applications

The Protein Journal - The extracellular enzyme with oxidase function was extracted from the Neonothopanus nambi luminescent fungus by using mild processing of mycelium with β-glucosidase and...     

Reusable System for Phenol Detection in an Aqueous Medium Based on Nanodiamonds and Extracellular Oxidase from Basidiomycete Neonothopanus nambi

Doklady Biochemistry and Biophysics - A reusable system for phenol determination in an aqueous medium was obtained by adsorption of extracellular oxidase from fungus Neonothopanus nambi onto...     

Downregulation of Serotonergic Gene Expression in the Raphe Nuclei of the Midbrain Under Chronic Social Defeat Stress in Male Mice

There is ample experimental evidence supporting the hypothesis that the brain serotonergic system is involved in the control of chronic social defeat stress (CSDS), depression, and anxiety. The study aimed to analyze mRNA levels of the serotonergic genes in the raphe nuclei of midbrain that may be associated with chronic social defeats consistently shown by male mice in special experimental settings. The serotonergic genes were the Tph2, Sert, Maoa, and Htr1a. The Bdnf and Creb genes were also studied. The experimental groups were composed of male mice with experience of defeats in 21 daily encounters and male mice with the same track record of defeats followed by a no-defeat period without agonistic interactions (relative rest for 14 days). It has been shown that mRNA levels of the Tph2, Maoa, Sert, Htr1a, Bdnf, and Creb genes in the raphe nuclei of defeated mice are decreased as compared with the controls. The expression of the serotonergic genes as well as the Creb gene is not restored to the control level after the 2 weeks of relative rest. mRNA levels of Bdnf gene are not recovered to the control levels, although some upregulation was observed in rested losers. CSDS experience inducing the development of mixed anxiety/depression-like state in male mice downregulates the expression of serotonergic genes associated with the synthesis, inactivation, and reception of serotonin. The Bdnf and Creb genes in the midbrain raphe nuclei are also downregulated under CSDS. Period of relative rest is not enough for most serotonergic genes to recover expression to the control levels.     

PARTAKE Survey of Public Knowledge and Perceptions of Clinical Research in India

Background

A public that is an informed partner in clinical research is important for ethical, methodological, and operational reasons. There are indications that the public is unaware or misinformed, and not sufficiently engaged in clinical research but studies on the topic are lacking. PARTAKE – Public Awareness of Research for Therapeutic Advancements through Knowledge and Empowerment is a program aimed at increasing public awareness and partnership in clinical research. The PARTAKE Survey is a component of the program.

Objective

To study public knowledge and perceptions of clinical research.

Methods

A 40-item questionnaire combining multiple-choice and open-ended questions was administered to 175 English- or Hindi-speaking individuals in 8 public locations representing various socioeconomic strata in New Delhi, India.

Results

Interviewees were 18–84 old (mean: 39.6, SD±16.6), 23.6% female, 68.6% employed, 7.3% illiterate, 26.3% had heard of research, 2.9% had participated and 58.9% expressed willingness to participate in clinical research. The following perceptions were reported (% true/% false/% not aware): ‘research benefits society’ (94.1%/3.5%/2.3%), ‘the government protects against unethical clinical research’ (56.7%/26.3%/16.9%), ‘research hospitals provide better care’ (67.2%/8.7%/23.9%), ‘confidentiality is adequately protected’ (54.1%/12.3%/33.5%), ‘participation in research is voluntary’ (85.3%/5.8%/8.7%); ‘participants treated like ‘guinea pigs’’ (20.7%/53.2%/26.0%), and ‘compensation for participation is adequate’ (24.7%/12.9%/62.3%).

Conclusions

Results suggest the Indian public is aware of some key features of clinical research (e.g., purpose, value, voluntary nature of participation), and supports clinical research in general but is unaware of other key features (e.g., compensation, confidentiality, protection of human participants) and exhibits some distrust in the conduct and reporting of clinical trials. Larger, cross-cultural surveys are required to inform educational programs addressing these issues.     

Structural and energetic determinants of primary proton transfer in bacteriorhodopsin.

In the light-driven bacteriorhodopsin proton pump, the first proton transfer step is from the retinal Schiff base to a nearby carboxylate group. The mechanism of this transfer step is highly controversial, in particular whether a direct proton jump is allowed. Here, we review the structural and energetic determinants of the direct proton transfer path computed by using a combined quantum mechanical/molecular mechanical approach. Both protein flexibility and electrostatic interactions play an important role in shaping the proton transfer energy profile. Detailed analysis of the energetics of putative transitions in the first half of the photocycle focuses on two elements that determine the likelihood that a given configuration of the active site is populated during the proton-pumping cycle. First, the rate-limiting barrier for proton transfer must be consistent with the kinetics of the photocycle. Second, the active-site configuration must be compatible with a productive overall pumping cycle.