Multiple chaperonins in bacteria—novel functions and non-canonical behaviors

Chaperonins are a class of molecular chaperones that assemble into a large double ring architecture with each ring constituting seven to nine subunits and enclosing a cavity for substrate encapsulation. The well-studied Escherichia coli chaperonin GroEL binds non-native substrates and encapsulates them in the cavity thereby sequestering the substrates from unfavorable conditions and allowing the substrates to fold. Using this mechanism, GroEL assists folding of about 10–15 % of cellular proteins. Surprisingly, about 30 % of the bacteria express multiple chaperonin genes. The presence of multiple chaperonins raises questions on whether they increase general chaperoning ability in the cell or have developed specific novel cellular roles. Although the latter view is widely supported, evidence for the former is beginning to appear. Some of these chaperonins can functionally replace GroEL in E. coli and are generally indispensable, while others are ineffective and likewise are dispensable. Additionally, moonlighting functions for several chaperonins have been demonstrated, indicating a functional diversity among the chaperonins. Furthermore, proteomic studies have identified diverse substrate pools for multiple chaperonins. We review the current perception on multiple chaperonins and their physiological and functional specificities.     

ProViDE: A software tool for accurate estimation of viral diversity in metagenomic samples

Given the absence of universal marker genes in the viral kingdom, researchers typically use BLAST (with stringent E-values) for taxonomic classification of viral metagenomic sequences. Since majority of metagenomic sequences originate from hitherto unknown viral groups, using stringent e-values results in most sequences remaining unclassified. Furthermore, using less stringent e-values results in a high number of incorrect taxonomic assignments. The SOrt-ITEMS algorithm provides an approach to address the above issues. Based on alignment parameters, SOrt-ITEMS follows an elaborate work-flow for assigning reads originating from hitherto unknown archaeal/bacterial genomes. In SOrt-ITEMS, alignment parameter thresholds were generated by observing patterns of sequence divergence within and across various taxonomic groups belonging to bacterial and archaeal kingdoms. However, many taxonomic groups within the viral kingdom lack a typical Linnean-like taxonomic hierarchy. In this paper, we present ProViDE (Program for Viral Diversity Estimation), an algorithm that uses a customized set of alignment parameter thresholds, specifically suited for viral metagenomic sequences. These thresholds capture the pattern of sequence divergence and the non-uniform taxonomic hierarchy observed within/across various taxonomic groups of the viral kingdom. Validation results indicate that the percentage of 'correct' assignments by ProViDE is around 1.7 to 3 times higher than that by the widely used similarity based method MEGAN. The misclassification rate of ProViDE is around 3 to 19% (as compared to 5 to 42% by MEGAN) indicating significantly better assignment accuracy. ProViDE software and a supplementary file (containing supplementary figures and tables referred to in this article) is available for download from http://metagenomics.atc.tcs.com/binning/ProViDE/     

Comparative shotgun proteomic analysis of Clostridium acetobutylicum from butanol fermentation using glucose and xylose

Background

Cytokinin is a plant hormone that plays a crucial role in several processes of plant growth and development. In recent years, major breakthroughs have been achieved in the elucidation of the metabolism, the signal perception and transduction, as well as the biological functions of cytokinin. An important activity of cytokinin is the involvement in chloroplast development and function. Although this biological function has already been known for 50 years, the exact mechanisms remain elusive.

Results

To elucidate the effects of altered endogenous cytokinin content on the structure and function of the chloroplasts, chloroplast subfractions (stroma and thylakoids) from transgenic Pssu-ipt and 35S:CKX1 tobacco (Nicotiana tabacum) plants with, respectively, elevated and reduced endogenous cytokinin content were analysed using two different 2-DE approaches. Firstly, thykaloids were analysed by blue-native polyacrylamide gel electrophoresis followed by SDS-PAGE (BN/SDS-PAGE). Image analysis of the gel spot pattern thus obtained from thylakoids showed no substantial differences between wild-type and transgenic tobacco plants. Secondly, a quantitative DIGE analysis of CHAPS soluble proteins derived from chloroplast subfractions indicated significant gel spot abundance differences in the stroma fraction. Upon identification by MALDI-TOF/TOF mass spectrometry, these proteins could be assigned to the Calvin-Benson cycle and photoprotective mechanisms.

Conclusion

Taken together, presented proteomic data reveal that the constitutively altered cytokinin status of transgenic plants does not result in any qualitative changes in either stroma proteins or protein complexes of thylakoid membranes of fully developed chloroplasts, while few but significant quantitative differences are observed in stroma proteins.     

Protein localization: reach out and touch the forespore

Bacterial proteins are typically sorted to subcellular regions with distinct physical characteristics that serve as cellular 'addresses', but many proteins are evidently sorted to specific areas that lack any apparent unique identity. Recent work in Bacillus subtilis suggests that such proteins may be localized by interacting with extracellular domains of proteins in an adjacent cellular compartment.     

Common binding site for disialyllactose and tri-peptide in C-fragment of tetanus neurotoxin

Clostridial neurotoxins are comprised of botulinum (BoNT) and tetanus (TeNT), which share significant structural and functional similarity. Crystal structures of the binding domain of TeNT complexed with disialyllactose (DiSia) and a tri-peptide Tyr-Glu-Trp (YEW) have been determined to 2.3 and 2.2 A, respectively. Both DiSia and YEW bind in a shallow cleft region on the surface of the molecule in the beta-trefoil domain, interacting with a set of common residues, Asp1147, Asp1214, Asn1216, and Arg1226. DiSia and YEW binding at the same site in tetanus toxin provides a putative site that could be occupied either by a ganglioside moiety or a peptide. Soaking experiments with a mixture of YEW and DiSia show that YEW competes with DiSia, suggesting that YEW can be used to block ganglioside binding. A comparison with the TeNT binding domain in complex with small molecules, BoNT/A and /B, provides insight into the different modes of ganglioside binding.     

A synonymous mutation in Yersinia enterocolitica yopE affects the function of the YopE type III secretion signal

Yersinia spp. inject virulence proteins called Yops into the cytosol of target eukaryotic cells in an effort to evade phagocytic killing via a dedicated protein-sorting pathway termed type III secretion. Previous studies have proposed that, unlike other protein translocation mechanisms, Yops are not recognized as substrates for secretion via a solely proteinaceous signal. Rather, at least some of this information may be encoded within yop mRNA. Herein, we report that the first seven codons of yopE, when fused to the reporter protein neomycin phosphotransferase (Npt), are sufficient for the secretion of YopE1-7-Npt when type III secretion is induced in vitro. Systematic mutagenesis of yopE codons 1 to 7 reveals that, like yopQ, codons 2, 3, 5, and 7 are sensitive to mutagenesis, thereby defining the first empirical similarity between the secretion signals of two type III secreted substrates. Like that of yopQ, the secretion signal of yopE exhibits a bipartite nature. This is manifested by the ability of codons 8 to 15 to suppress point mutations in the minimal secretion signal that change the amino acid specificities of particular codons or that induce alterations in the reading frame. Further, we have identified a single nucleotide position in codon 3 that, when mutated, conserves the predicted amino acid sequence of the YopE1-7-Npt but abrogates secretion of the reporter protein. When introduced into the context of the full-length yopE gene, the single-nucleotide mutation reduces the type III injection of YopE into HeLa cells, even though the predicted amino acid sequence remains the same. Thus, yopE mRNA appears to encode a property that mediates the type III injection of YopE.     

The population structure of African cultivated rice oryza glaberrima (Steud.): evidence for elevated levels of linkage disequilibrium caused by admixture with O. sativa and ecological adaptation

Genome-wide linkage disequilibrium (LD) was investigated for 198 accessions of Oryza glaberrima using 93 nuclear microsatellite markers. Significantly elevated levels of LD were detected, even among distantly located markers. Free recombination among loci at the population genetic level was shown (1) by a lack of decay in LD among markers on the same chromosome and (2) by a strictly increasing composite likelihood function for the recombination parameter. This suggested that the elevation in LD was due not to physical linkage but to other factors, such as population structure. A Bayesian clustering analysis confirmed this hypothesis, indicating that the sample of O. glaberrima in this study was subdivided into at least five cryptic subpopulations. Two of these subpopulations clustered with control samples of O. sativa, subspecies indica and japonica, indicating that some O. glaberrima accessions represent admixtures. The remaining three O. glaberrima subpopulations were significantly associated with specific combinations of phenotypic traits-possibly reflecting ecological adaptation to different growing environments.     

Age-associated decreased activities of mitochondrial electron transport chain complexes in heart and skeletal muscle: role of L-carnitine

The mitochondrial respiratory chain is a powerful source of reactive oxygen species (ROS), considered as the pathogenic agent of many diseases and aging. L-Carnitine (4-N-trimethylammonium-3-hydroxybutric acid) plays an important role in transport of fatty acid from cytoplasm to mitochondria for energy production. Previous studies in our laboratory reported L-carnitine as a free radical scavenger in aged rats. In the present study we focused the effect of L-carnitine on the activities of electron transport chain in young and aged rats. The activities of electron transport chain complexes were found to be significantly decreased in aged rats when compared to young control rats. Supplementation of carnitine to young and aged rats for 14 and 21 days improved the electron transport chain complexes levels in aged rats when compared with young rats in duration dependent manner. No significant changes were observed in young rats. Our result suggested that L-carnitine improved the activities of electron transport chain enzymes there by improving the energy status in aged rats.