Unusual routes of protein secretion: the easy way out

Increasing numbers of polypeptides are being discovered that lack a cleavable hydrophobic signal sequence and are released from cells without passing through the classical secretory pathway. This article reviews the current knowledge of these alternative secretion pathways in prokaryotes and eukaryotes and discusses whether the mechanisms described in bacteria and yeast can be used as paradigms to explain unusual secretory phenomena in animal cells.     

An easy way to reduce PIT-tag loss in rodents

Passive integrated transponders (PIT-tags) have been shown to be a reliable method for individually marking rodents in field studies. The loss rates of PIT-tags vary, depending on the species, from 3.6 to 7.2%. We used PIT-tags to individually mark a population of edible dormice. Due to a relatively high loss rate of transponders in the first year (7.4%) we used a tissue adhesive to seal the punctuation wound in the second year. With this method we could reduce the tag-loss rate to 2.2%. We therefore recommend using a tissue adhesive, which is easy and fast to apply, after injecting a transponder.     

Estimating total genic diversity in the house mouse

In a survey of variation in both electrophoretic charge and thermostability at 14 structural loci in 40 strains of Mus musculus, 27 electromorphs (polypeptides differing in electrophoretic charge) and 20 thermomorphs (polypeptides differing in thermostability) were found. Electrophoresis detected 11 new variants within thermomorphs, and the heat denaturation technique detected four new variants within electromorphs. From these data, and making the assumption that both techniques are independent of each other, it is estimated that the actual total number of alleles at the 14 loci is 53, or an average of 3.79 per locus (1.96 per electromorph), and that electrophoresis apparently detects one-third of the variants, thus describing about 50% of the alleles at structural gene loci in the house mouse.     

Estimating and comparing the diversity of marine actinobacteria

This paper reviews the application of species richness estimators to microbial diversity data and describes phylogenetic approaches to comparing microbial communities. The techniques are demonstrated using a community of marine actinobacteria. Results demonstrate that marine environments harbour massive actinobacterial diversity. Furthermore, these predictions are likely to be severe underestimates due to the use of arbitrary OTU definitions.     

White gels: an easy way to preserve methylene blue stained gels

Methylene blue can be used as a stain for visualizing nucleic acids in polyacrylamide gel electrophoresis. However, its relatively low sensitivity and reversible binding make it a temporary stain that diffuses from the gel relatively fast. Here we describe a very simple method for fixing methylene blue bands in nucleic acid polyacrylamide gels. The procedure makes the methylene blue stain permanent and increases the visibility of the bands, also contributing to increasing the sensitivity of methylene blue.     

An easy way to include weak alignment constraints into NMR structure calculations

We have recently shown that an energy penalty for the incorporation of residual tensorial constraints into molecular structure calculations can be formulated without the explicit knowledge of the Saupe orientation tensor (Moltke and Grzesiek, J. Biomol. NMR, 1999, 15, 77–82). Here we report the implementation of such an algorithm into the program X-PLOR. The new algorithm is easy to use and has good convergence properties. The algorithm is used for the structure refinement of the HIV-1 Nef protein using 252 dipolar coupling restraints. The approach is compared to the conventional penalty function with explicit knowledge of the orientation tensor's amplitude and rhombicity. No significant differences are found with respect to speed, Ramachandran core quality or coordinate precision.     

Estimating the diversity of peptide populations from limited sequence data

MOTIVATION: Combinatorial libraries of peptides such as those displayed on the surface of a bacteriophage particle have become widely used tools for characterizing protein-protein and protein-small molecule interactions. The quality of a library frequently depends on its completeness, or diversity-the proportion of possible sequences actually present in the library. The diversity of these libraries is frequently quoted on the basis of phage titers that provide little information about their completeness. RESULTS: Here, an analytical expression for diversity is introduced and a method for estimating the diversity of a peptide library from the sequences of a limited number of the members of the library is demonstrated. The diversities of a number of computationally constructed and actual peptide libraries are estimated using this method.     

Estimating the contribution of mutation, recombination and gene conversion in the generation of haplotypic diversity

Recombination occurs through both homologous crossing over and homologous gene conversion during meiosis. The contribution of recombination relative to mutation is expected to be dramatically reduced in inbreeding organisms. We report coalescent-based estimates of the recombination parameter (rho) relative to estimates of the mutation parameter (theta) for 18 genes from the highly self-fertilizing grass, wild barley, Hordeum vulgare ssp. spontaneum. Estimates of rho/theta are much greater than expected, with a mean rho/theta approximately 1.5, similar to estimates from outcrossing species. We also estimate rho with and without the contribution of gene conversion. Genotyping errors can mimic the effect of gene conversion, upwardly biasing estimates of the role of conversion. Thus we report a novel method for identifying genotyping errors in nucleotide sequence data sets. We show that there is evidence for gene conversion in many large nucleotide sequence data sets including our data that have been purged of all detectable sequencing errors and in data sets from Drosophila melanogaster, D. simulans, and Zea mays. In total, 13 of 27 loci show evidence of gene conversion. For these loci, gene conversion is estimated to contribute an average of twice as much as crossing over to total recombination.     

Estimating species richness: still a long way off!

Xu et al., in this issue of the Journal of Vegetation Science, compare several species richness estimators. All the non‐parametric estimators, such as Chao and jackknife estimators, underestimated the true number, whereas all the area‐based models, based on species–area curves, overestimated it. No reliable method yet exists to predict the number of species in an area that is appreciably larger than the one(s) sampled.     

The pEAQ vector series: the easy and quick way to produce recombinant proteins in plants

Promoter regions of six sugarcane Loading Stem Gene (ScLSG) alleles were analyzed using bioinformatic and transgenic approaches. Stable transgene expression analyses, on multiple independent lines per construct, revealed differences between ScLSG promoters in absolute levels and in tissue-selectivity of luciferase reporter activity. Four promoters drove peak expression in the sucrose-loading zone and maintained substantial expression throughout mature stems. One drove a pattern of gradual increase along the stem maturation profile. In general, stem: root expression ratio increased with plant age. The ScLSG5 promoter had the fewest light-enhanced and root-expression motifs in bioinformatic analysis, and drove the highest level and specificity of transgene expression in stems. This indicates the potential to further improve the stem specificity of ScLSG promoter sequences by eliminating enhancers of expression in other tissues. An intron in the 5′UTR was important for expression strength. The ScLSG promoters will be useful for research and biotechnology in sugarcane, where the tailored expression of transgenes in stems is important for enhanced accumulation of sugar or value-added products, and for development as a bioenergy feedstock.     

Tick salivary proteins offer the lyme disease spirochetes an easy ride and another way to hide

The ability of the Lyme disease spirochetes to establish an infection in mammals is dependent in part on proteins of tick origin. Schuijt et?al. (2011) investigate the role of the tick-derived protein, TSLPI, in spirochete transmission and in the evasion of killing by the lectin complement pathway.