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.     

Genome-wide differentially methylated genes in prostate cancer tissues from African-American and Caucasian men

Increasing evidence suggests that aberrant DNA methylation changes may contribute to prostate cancer (PCa) ethnic disparity. To comprehensively identify DNA methylation alterations in PCa disparity, we used the Illumina 450K methylation platform to interrogate the methylation status of 485,577 CpG sites focusing on gene-associated regions of the human genome. Genomic DNA from African-American (AA; 7 normal and 3 cancers) and Caucasian (Cau; 8 normal and 3 cancers) was used in the analysis. Hierarchical clustering analysis identified probe-sets unique to AA and Cau samples, as well as common to both. We selected 25 promoter-associated novel CpG sites most differentially methylated by race (fold change > 1.5-fold; adjusted P < 0.05) and compared the β-value of these sites provided by the Illumina, Inc. array with quantitative methylation obtained by pyrosequencing in 7 prostate cell lines. We found very good concordance of the methylation levels between β-value and pyrosequencing. Gene expression analysis using qRT-PCR in a subset of 8 genes after treatment with 5-aza-2′-deoxycytidine and/or trichostatin showed up-regulation of gene expression in PCa cells. Quantitative analysis of 4 genes, SNRPN, SHANK2, MST1R, and ABCG5, in matched normal and PCa tissues derived from AA and Cau PCa patients demonstrated differential promoter methylation and concomitant differences in mRNA expression in prostate tissues from AA vs. Cau. Regression analysis in normal and PCa tissues as a function of race showed significantly higher methylation prevalence for SNRPN (P = 0.012), MST1R (P = 0.038), and ABCG5 (P < 0.0002) for AA vs. Cau samples. We selected the ABCG5 and SNRPN genes and verified their biological functions by Western blot analysis and siRNA gene knockout effects on cell proliferation and invasion in 4 PCa cell lines (2 AA and 2 Cau patients-derived lines). Knockdown of either ABCG5 or SNRPN resulted in a significant decrease in both invasion and proliferation in Cau PCa cell lines but we did not observe these remarkable loss-of-function effects in AA PCa cell lines. Our study demonstrates how differential genome-wide DNA methylation levels influence gene expression and biological functions in AA and Cau PCa.     

Inhibition of Listeria innocua and Brochothrix thermosphacta in vacuum-packaged meat by addition of bacteriocinogenic Lactobacillus curvatus CRL705 and its bacteriocins

AIMS: To evaluate the inhibition effectiveness of Lactobacillus curvatus CRL705 used as a bioprotective culture and of its bacteriocins, lactocin 705 and lactocin AL705, against Listeria innocua, Brochothrix thermosphacta and indigenous lactic acid bacteria (LAB) in vacuum-packaged meat stored at 2 degrees C. METHODS AND RESULTS: The live culture of Lact. curvatus CRL705 as well as synthetic lactocin 705 and purified lactocin AL705 were shown to be similarly effective in preventing the growth of B. thermosphacta and L. innocua in meat discs in contrast to control samples in which these micro-organisms grew rapidly, their numbers increasing by 3.0- and 2.1-log cycles respectively. In addition, indigenous LAB population showed a lower growth rate in the presence of lactocin 705. Bacteriocin activity was detected in the meat discs during 36 days at 2 degrees C irrespective of the biopreservation strategy applied. Changes in pH were not significantly different in meat discs treated with the protective culture when compared with control samples. CONCLUSIONS: Lactobacillus curvatus CRL705 and the produced bacteriocins, lactocin 705 and lactocin AL 705, were effective in inhibiting L. innocua and B. thermosphacta. The use of the bioprotective culture in refrigerated vacuum-packaged fresh meat would be more feasible from an economic and legal point of view. SIGNIFICANCE AND IMPACT OF THE STUDY: Establishment of biopreservation as a method to ensure the microbiological safety of vacuum-packaged fresh meat at 2 degrees C.     

Comparison of classification methods for detecting associations between SNPs and chick mortality

Multi-category classification methods were used to detect SNP-mortality associations in broilers. The objective was to select a subset of whole genome SNPs associated with chick mortality. This was done by categorizing mortality rates and using a filter-wrapper feature selection procedure in each of the classification methods evaluated. Different numbers of categories (2, 3, 4, 5 and 10) and three classification algorithms (naïve Bayes classifiers, Bayesian networks and neural networks) were compared, using early and late chick mortality rates in low and high hygiene environments. Evaluation of SNPs selected by each classification method was done by predicted residual sum of squares and a significance test-related metric. A naïve Bayes classifier, coupled with discretization into two or three categories generated the SNP subset with greatest predictive ability. Further, an alternative categorization scheme, which used only two extreme portions of the empirical distribution of mortality rates, was considered. This scheme selected SNPs with greater predictive ability than those chosen by the methods described previously. Use of extreme samples seems to enhance the ability of feature selection procedures to select influential SNPs in genetic association studies.     

Structural studies on zinc(II) complexes with 2-benzoylpyridine-derived hydrazones

2-Benzoylpyridine-phenylhydrazone (H2BzPh), 2-benzoylpyridine-para-chloro-phenylhydrazone (H2BzpClPh), and 2-benzoylpyridine-para-nitro-phenyl (H2BzpNO₂Ph) hydrazone were obtained and fully characterized, as well as their zinc(II) complexes [Zn(H2BzPh)Cl₂] (1), [Zn(H2BzClPh)Cl₂] (2) and [Zn(H2BzpNO₂Ph)Cl₂] (3). During the syntheses of complex 1 a second product crystallized, which was characterized as [Zn(2BzPh)₂] (1a). Upon re-crystallization in 1:9 DMSO:acetone conversion of 2 into [Zn(H2BzpClPh)Cl₂] · H₂O (2a) and of 3 into [Zn(2BzpNO₂Ph)Cl(DMSO)] (3a) occurred. The crystal structures of 1a, 2a and 3a were determined. In 1a the two nearly perpendicular H2BzPh ligands give rise to a distorted octahedral environment around the metal. The 5-fold coordination around the metal is completed with two chloride ions in 2a and with one chloride and one oxygen atom from DMSO in 3a.     

Adverse environmental conditions influence age-related innate immune responsiveness

Background-  

The innate immune system plays an important role in the recognition and induction of protective responses against infectious pathogens, whilst there is increasing evidence for a role in mediating chronic inflammatory diseases at older age. Despite indications that environmental conditions can influence the senescence process of the adaptive immune system, it is not known whether the same holds true for the innate immune system. Therefore we studied whether age-related innate immune responses are similar or differ between populations living under very diverse environmental conditions.