Using chemical derivatization and mass spectrometric analysis to characterize the post-translationally modified Staphylococcus aureus surface protein G

The Staphylococcus aureus surface protein G (SasG) is an important mediator of biofilm formation in virulent S. aureus strains. A detailed analysis of its primary sequence has not been reported to date. SasG is highly abundant in the cell wall of the vancomycin-intermediate S. aureus strain HIP5827, and was purified and subjected to sequence analysis by MS. Data from MALDI-TOF and LC-MS/MS experiments confirmed the predicted N-terminal signal peptide cleavage site at residue A₅₁ and the C-terminal cell wall anchor site at residue T₁₀₈₆. The protein was also derivatized with N-succinimidyloxycarbonyl-methyl-tris(2,4,6-trimethoxyphenyl) phosphonium bromide (TMPP-Ac-OSu) to assess the presence of additional N-terminal sites of mature SasG. TMPP-derivatized SasG peptides featured m/z peaks with a 572 Da mass increase over the equivalent underivatized peptides. Multiple N-terminal peptides, all of which were observed in the 150 amino acid segment following the signal peptide cleavage at the residue A₅₁, were characterized from MS and MS/MS data, suggesting a series of successive N-terminal truncations of SasG. A strategy combining TMPP derivatization, multiple enzyme digestions to generate overlapping peptides and detailed MS analysis will be useful to determine and understand functional implications of PTMs in bacterial cell wall-anchored proteins, which are frequently involved in the modulation of virulence-associated bacterial surface properties.     

The contribution of maximal force production to explosive movement among young collegiate athletes

Critical to multidimensional sport conditioning is a systematic knowledge of the interactions between fitness components, as well as the transference relationships to performance. The purpose of this investigation was to examine the relationships between lower body muscular strength and several fundamental explosive performance measures. Fifty-four men and women collegiate athletes were tested to determine (a) lower-body muscular strength (1 repetition maximum barbell back squat), (b) countermovement vertical jump height and peak power output, (c) standing broad jump distance, (d) agility (cone T-test time), (e) sprint acceleration (m.s(-2)), and (f) sprint velocity (m.s(-1)). Analyses were performed using Pearson r correlations to examine these relationships. Partial correlations tested for relationships between performance measures while controlling for muscular strength. T-tests were performed to assess the difference between men and women. Correlation data demonstrated that significant (p < 0.01) strong linear relationships were indicated between muscular strength and power, as well as every sport-performance field tests. However, when controlling for strength with partial correlation, each of these relationships appreciably diminished. Significant differences (p < 0.05) were found between men and women for each of the performance tests. Muscular strength, peak power output, vertical jumping ability, standing broad jump, agility, sprint acceleration, and sprint velocity were all shown to be very highly related. Further examination demonstrated that body mass-adjusted muscular strength is more highly related to performance measures than is absolute muscular strength. Current correlation data provide a quantified look at the interaction between muscular fitness components, as well as the transfer relationship to several athletic-specific performance measures.     

Grasses through space and time: An overview of the biogeographical and macroevolutionary history of Poaceae

Grasses are widespread on every continent and are found in all terrestrial biomes. The dominance and spread of grasses and grassland ecosystems have led to significant changes in Earth′s climate, geochemistry, and biodiversity. The abundance of DNA sequence data, particularly chloroplast sequences, and advances in placing grass fossils within the family allows for a reappraisal of the family′s origins, timing, and geographic spread and the factors that have promoted diversification. We reconstructed a time-calibrated grass phylogeny and inferred ancestral areas using chloroplast DNA sequences from nearly 90% of extant grass genera. With a few notable exceptions, the phylogeny is well resolved to the subtribal level. The family began to diversify in the Early–Late Cretaceous (crown age of 98.54 Ma) on West Gondwana before the complete split between Africa and South America. Vicariance from the splitting of Gondwana may be responsible for the initial divergence in the family. However, Africa clearly served as the center of origin for much of the early diversification of the family. With this phylogenetic, temporal, and spatial framework, we review the evolution and biogeography of the family with the aim to facilitate the testing of biogeographical hypotheses about its origins, evolutionary tempo, and diversification. The current classification of the family is discussed with an extensive review of the extant diversity and distribution of species, molecular and morphological evidence supporting the current classification scheme, and the evidence informing our understanding of the biogeographical history of the family.     

Modeling the influence of the extrinsic musculature on phonation

Biomechanics and Modeling in Mechanobiology - Neck muscles play important roles in various physiological tasks, including swallowing, head stabilization, and phonation. The mechanisms by which neck...     

Avian eggshell thickness in relation to egg morphometrics,embryonic development,and mercury contamination

Eggshell thickness is important for physiological, ecological, and ecotoxicological studies on birds; however, empirical eggshell thickness measurements for many species and regions are limited. We measured eggshell thickness at the equator and the egg poles for 12 avian species and related eggshell thickness to egg morphometrics, embryonic development, egg status, and mercury contamination. Within an egg, eggshells were approximately 5.1% thicker at the equator than the sharp pole of the egg, although this difference varied among species (0.6%–9.8%). Within Forster's tern (Sterna forsteri), where eggshell thickness was measured at 5 equally spaced positions along the longitude of the egg, eggshell thickness changed more rapidly near the sharp pole of the egg compared to near the blunt pole of the egg. Within species, eggshell thickness was related to egg width and egg volume for six of the 12 species but was not related to egg length for any species. Among species, mean eggshell thickness was strongly related to species mean egg width, egg length, egg volume, and bird body mass, although species mean body mass was the strongest predictor of species mean eggshell thickness. Using three species (American avocet [Recurvirostra americana], black‐necked stilt [Himantopus mexicanus], and Forster's tern), whose nests were carefully monitored, eggshell thickness (including the eggshell membrane) did not differ among viable, naturally abandoned, dead, or failed‐to‐hatch eggs; was not related to total mercury concentrations of the egg content; and did not decrease with embryonic age. Our study also provides a review of all existing eggshell thickness data for these 12 species.     

Gene disruption by structural mutations drives selection in US rice breeding over the last century

The genetic basis of general plant vigor is of major interest to food producers, yet the trait is recalcitrant to genetic mapping because of the number of loci involved, their small effects, and linkage. Observations of heterosis in many crops suggests that recessive, malfunctioning versions of genes are a major cause of poor performance, yet we have little information on the mutational spectrum underlying these disruptions. To address this question, we generated a long-read assembly of a tropical japonica rice (Oryza sativa) variety, Carolina Gold, which allowed us to identify structural mutations (>50 bp) and orient them with respect to their ancestral state using the outgroup, Oryza glaberrima. Supporting prior work, we find substantial genome expansion in the sativa branch. While transposable elements (TEs) account for the largest share of size variation, the majority of events are not directly TE-mediated. Tandem duplications are the most common source of insertions and are highly enriched among 50-200bp mutations. To explore the relative impact of various mutational classes on crop fitness, we then track these structural events over the last century of US rice improvement using 101 resequenced varieties. Within this material, a pattern of temporary hybridization between medium and long-grain varieties was followed by recent divergence. During this long-term selection, structural mutations that impact gene exons have been removed at a greater rate than intronic indels and single-nucleotide mutations. These results support the use of ab initio estimates of mutational burden, based on structural data, as an orthogonal predictor in genomic selection.     

CD4(+) T cells and gamma interferon in the long-term control of persistent friend retrovirus infection

We have used the Friend virus model to determine the basic mechanisms by which the immune system can control persistent retroviral infections. Previously we showed that CD4(+) T cells play an essential role in keeping persistent retrovirus in check. The present in vitro experiments with a Friend virus-specific CD4(+) T-cell clone revealed that these cells produce gamma interferon (IFN-gamma), which acts with two distinct mechanisms of antiviral activity. First, IFN-gamma had a direct inhibitory effect on virus production. This inhibitory effect was noncytolytic and, interestingly, was not associated with decreased cell surface expression of viral antigens. The second mechanism of IFN-gamma-mediated antiviral activity was an enhancement of CD4(+) T-cell-mediated cytolytic activity. We also found an in vivo role for IFN-gamma in the control of persistent Friend virus infections. Neutralization of IFN-gamma in persistently infected mice resulted in significantly increased levels of virus in the spleen, and a significant percentage of IFN-gamma-deficient mice were unable to maintain long-term control over Friend virus infections.     

Crystal structure of inhibitor-bound P450BM-3 reveals open conformation of substrate access channel

P450BM-3 is an extensively studied P450 cytochrome that is naturally fused to a cytochrome P450 reductase domain. Crystal structures of the heme domain of this enzyme have previously generated many insights into features of P450 structure, substrate binding specificity, and conformational changes that occur on substrate binding. Although many P450s are inhibited by imidazole, this compound does not effectively inhibit P450BM-3. Omega-imidazolyl fatty acids have previously been found to be weak inhibitors of the enzyme and show some unusual cooperativity with the substrate lauric acid. We set out to improve the properties of these inhibitors by attaching the omega-imidazolyl fatty acid to the nitrogen of an amino acid group, a tactic that we used previously to increase the potency of substrates. The resulting inhibitors were significantly more potent than their parent compounds lacking the amino acid group. A crystal structure of one of the new inhibitors bound to the heme domain of P450BM-3 reveals that the mode of interaction of the amino acid group with the enzyme is different from that previously observed for acyl amino acid substrates. Further, required movements of residues in the active site to accommodate the imidazole group provide an explanation for the low affinity of imidazole itself. Finally, the previously observed cooperativity with lauric acid is explained by a surprisingly open substrate-access channel lined with hydrophobic residues that could potentially accommodate lauric acid in addition to the inhibitor itself.