Quantitative analysis of complex protein mixtures using isotope-coded affinity tags.

We describe an approach for the accurate quantification and concurrent sequence identification of the individual proteins within complex mixtures. The method is based on a class of new chemical reagents termed isotope-coded affinity tags (ICATs) and tandem mass spectrometry. Using this strategy, we compared protein expression in the yeast Saccharomyces cerevisiae, using either ethanol or galactose as a carbon source. The measured differences in protein expression correlated with known yeast metabolic function under glucose-repressed conditions. The method is redundant if multiple cysteinyl residues are present, and the relative quantification is highly accurate because it is based on stable isotope dilution techniques. The ICAT approach should provide a widely applicable means to compare quantitatively global protein expression in cells and tissues.     

CGRP 27-37 analogues with high affinity to the CGRP1 receptor show antagonistic properties in a rat blood flow assay

CGRP Y0-28-37 is known as a selective CGRP1 receptor antagonist. We succeeded in optimising the CGRP1 receptor affinity of this fragment by multiple amino acid replacement. The analogues [p34, F35]CGRP 27-37 and [D31, p34, F35]CGRP 27-37 exhibit a 100-fold increased affinity compared to the unmodified segment. Receptor binding studies were performed with human neuroblastoma cells SK-N-MC, which selectively express the hCGRP1 receptor. Blood flow, which is increased by exogenous CGRP, was measured in the right femoral artery. Preincubation of the rats with [p34, F35]CGRP 27-37 and [D31, p34, F35]CGRP 27-37 led to a significant decrease in CGRP induced increase in vascular conductance indicating the antagonistic properties of these compounds. Interestingly, an exchange of the amino acid Asn31 to Asp31 in [p34, F35]CGRP 27-37 shortened the period of the antagonistic effect significantly, suggestive of a different rate of metabolism for the two ligands. Secondary structure investigations obtained by circular dichroism measurements revealed that an increase in ordered structure correlates with high binding affinity.     

Single cell studies of the cell cycle and some models

Analysis of growth and division often involves measurements made on cell populations, which tend to average data. The value of single cell analysis needs to be appreciated, and models based on findings from single cells should be taken into greater consideration in our understanding of the way in which cell size and division are co-ordinated. Examples are given of some single cell analyses in mammalian cells, yeast and other microorganisms. There is also a short discussion on how far the results are in accord with simple models.     

Can Andean medicine coexist with biomedical healthcare? A comparison of two rural communities in Peru and Bolivia

ABSTRACT: BACKGROUND: It is commonly assumed that indigenous medical systems are strong in developing countries because biomedicine is physically inaccessible or financially not affordable. This paper compares the health-seeking behavior of households from rural Andean communities at a Peruvian and a Bolivian study site. The main research question was whether the increased presence of biomedicine led to a displacement of Andean indigenous medical practices or to coexistence of the two healing traditions. Methodology: Interviews were conducted between June 2006 and December 2008 with 18 households at each study site. Qualitative identification and analysis of households' therapeutic strategies and use of remedies was complemented by quantitative assessment of the incidence of culture-bound illnesses in local ethnobiological inventories. RESULTS: Our findings indicate that the health-seeking behavior of Andean households is independent of the degree of availability of biomedical facilities in terms of quality of services provided, physical accessibility, and financial affordability, except for specific practices such as childbirth. Preference for natural remedies over pharmaceuticals coexisted with biomedical healthcare that was both accessible and affordable. Furthermore, our results show that greater access to biomedicine does not lead to less prevalence of Andean indigenous medical knowledge, as represented by the levels of knowledge about culture-bound illnesses. CONCLUSIONS: The take-home lesson for health policy-makers from this study is that the main obstacle to improved use of biomedicine in resource-poor rural areas might not be infrastructural or economic alone. Rather, it may lie in lack of sufficient recognition by biomedical practitioners of the value and importance of indigenous medical systems. We propose that the implementation of health care in indigenous communities be designed as a process of joint development of complementary knowledge and practices from indigenous and biomedical health traditions.     

Methods for peptide identification by spectral comparison

Background  

Tandem mass spectrometry followed by database search is currently the predominant technology for peptide sequencing in shotgun proteomics experiments. Most methods compare experimentally observed spectra to the theoretical spectra predicted from the sequences in protein databases. There is a growing interest, however, in comparing unknown experimental spectra to a library of previously identified spectra. This approach has the advantage of taking into account instrument-dependent factors and peptide-specific differences in fragmentation probabilities. It is also computationally more efficient for high-throughput proteomics studies.     

Removing celiac disease-related gluten proteins from bread wheat while retaining technological properties: a study with Chinese Spring deletion lines

Background  

Gluten proteins can induce celiac disease (CD) in genetically susceptible individuals. In CD patients gluten-derived peptides are presented to the immune system, which leads to a CD4⁺ T-cell mediated immune response and inflammation of the small intestine. However, not all gluten proteins contain T-cell stimulatory epitopes. Gluten proteins are encoded by multigene loci present on chromosomes 1 and 6 of the three different genomes of hexaploid bread wheat (Triticum aestivum) (AABBDD).