Electromyographic patterns of individuals suffering from lateral tennis elbow.

This study investigated the applicability of using surface electromyography (EMG) as a tool for differentiating between persons suffering from lateral tennis elbow and the healthy age-matched adults. Temporal muscle activation patterns of the tennis elbow group were evaluated to determine if they varied between subject groups and if noted variations might be interpreted as arresting or exacerbating the injury. Sixteen subjects (Healthy Controls, n = 6; Tennis Elbow, n = 10) were tested under simulated tennis playing conditions. All subjects were males (Healthy group (CON) 38.8 +/- 13.1, Injured group (INJ) 40.8 +/- 10.8 yrs). EMG response data, temporal and spatial muscle activities, of the forearm extensors (Ext), the forearm flexors (Flex) and the triceps (Tri) were recorded for each subject during a single test session using all combinations of three different velocities on three different racket head impact locations. Data were collected at a frequency of 1000 Hz. Statistical analysis was performed using a 2 x 3 x 3 (Health status x Impact velocity x Impact location) ANOVA with repeated measures. Results indicated statistically significant differences (p < 0.05) between the CON and INJ subject groups for the response variables associated with forearm extensor muscle activation. During simulated play, the INJ group employed an earlier, longer, and greater activation of Ext than the CON group, such changes may be considered detrimental to the healing process. These results support the use of surface EMG to quantify differences in muscle activation strategies employed by individuals suffering from soft tissue muscle microtrauma injuries and healthy controls.     

An efficient strategy for assignment of cross-peaks in 3D heteronuclear NOESY experiments

The question is addressed of how maximal structural NOE data on double labelled proteins can be acquired with a minimal set of NOESY experiments. Two 3D-NOESY spectra are reported which, in concert with other commonly used spectra, provide a convenient strategy for NOE assignment. The 3D CNH-NOESY and 3D NCH-NOESY provide NOE connectivities between amide protons and carbon-bound protons and constitute orthogonal heteronuclear filters which eliminate diagonal signals, considerably improving spectral quality. Two different heteronuclear chemical shift dimensions are recorded in the spectra, thus exploiting the extra dispersion of the heteronucleus and considerably simplifying assignment.     

On the structure and operation of type I DNA restriction enzymes.

Type I DNA restriction enzymes are large, molecular machines possessing DNA methyltransferase, ATPase, DNA translocase and endonuclease activities. The ATPase, DNA translocase and endonuclease activities are specified by the restriction (R) subunit of the enzyme. We demonstrate that the R subunit of the Eco KI type I restriction enzyme comprises several different functional domains. An N-terminal domain contains an amino acid motif identical with that forming the catalytic site in simple restriction endonucleases, and changes within this motif lead to a loss of nuclease activity and abolish the restriction reaction. The central part of the R subunit contains amino acid sequences characteristic of DNA helicases. We demonstrate, using limited proteolysis of this subunit, that the helicase motifs are contained in two domains. Secondary structure prediction of these domains suggests a structure that is the same as the catalytic domains of DNA helicases of known structure. The C-terminal region of the R subunit can be removed by elastase treatment leaving a large fragment, stable in the presence of ATP, which can no longer bind to the other subunits of Eco KI suggesting that this domain is required for protein assembly. Considering these results and previous models of the methyltransferase part of these enzymes, a structural and operational model of a type I DNA restriction enzyme is presented.     

Posttranslational modifications of serine protease TMPRSS13 regulate zymogen activation,proteolytic activity,and cell surface localization

TMPRSS13, a member of the type II transmembrane serine protease (TTSP) family, harbors four N-linked glycosylation sites in its extracellular domain. Two of the glycosylated residues are located in the scavenger receptor cysteine-rich (SRCR) protein domain, while the remaining two sites are in the catalytic serine protease (SP) domain. In this study, we examined the role of N-linked glycosylation in the proteolytic activity, autoactivation, and cellular localization of TMPRSS13. Individual and combinatory site-directed mutagenesis of the glycosylated asparagine residues indicated that glycosylation of the SP domain is critical for TMPRSS13 autoactivation and catalytic activity toward one of its protein substrates, the prostasin zymogen. Additionally, SP domain glycosylation-deficient TMPRSS13 displayed impaired trafficking of TMPRSS13 to the cell surface, which correlated with increased retention in the endoplasmic reticulum. Importantly, we showed that N-linked glycosylation was a critical determinant for subsequent phosphorylation of endogenous TMPRSS13. Taken together, we conclude that glycosylation plays an important role in regulating TMPRSS13 activation and activity, phosphorylation, and cell surface localization.     

Mutation rate dynamics reflect ecological change in an emerging zoonotic pathogen

Mutation rates vary both within and between bacterial species, and understanding what drives this variation is essential for understanding the evolutionary dynamics of bacterial populations. In this study, we investigate two factors that are predicted to influence the mutation rate: ecology and genome size. We conducted mutation accumulation experiments on eight strains of the emerging zoonotic pathogen Streptococcus suis. Natural variation within this species allows us to compare tonsil carriage and invasive disease isolates, from both more and less pathogenic populations, with a wide range of genome sizes. We find that invasive disease isolates have repeatedly evolved mutation rates that are higher than those of closely related carriage isolates, regardless of variation in genome size. Independent of this variation in overall rate, we also observe a stronger bias towards G/C to A/T mutations in isolates from more pathogenic populations, whose genomes tend to be smaller and more AT-rich. Our results suggest that ecology is a stronger correlate of mutation rate than genome size over these timescales, and that transitions to invasive disease are consistently accompanied by rapid increases in mutation rate. These results shed light on the impact that ecology can have on the adaptive potential of bacterial pathogens.     

Ploidy and recombination proficiency shape the evolutionary adaptation to constitutive DNA replication stress

In haploid budding yeast, evolutionary adaptation to constitutive DNA replication stress alters three genome maintenance modules: DNA replication, the DNA damage checkpoint, and sister chromatid cohesion. We asked how these trajectories depend on genomic features by comparing the adaptation in three strains: haploids, diploids, and recombination deficient haploids. In all three, adaptation happens within 1000 generations at rates that are correlated with the initial fitness defect of the ancestors. Mutations in individual genes are selected at different frequencies in populations with different genomic features, but the benefits these mutations confer are similar in the three strains, and combinations of these mutations reproduce the fitness gains of evolved populations. Despite the differences in the selected mutations, adaptation targets the same three functional modules in strains with different genomic features, revealing a common evolutionary response to constitutive DNA replication stress.     

X-ray structures of new dipeptide taste ligands

The molecular basis of sweet taste was investigated by carrying out the crystal state conformational analysis by X-ray diffraction of the following dipeptide taste igands:N-3,3-dimethylbutyl-aspartyl-phenylalanine methyl ester, I (N-DMB-Asp-Phe-OMe), its sodium salt (N-DMB-Asp-Phe-ONa), II , aspartyl-D -2-aminobutyric acid-(S)-α-ethylbenzylamide, III (Asp-D -Abu-(S)-α-ethylbenzylamide), aspartyl-N′-((2,2,5,5-tetramethylcyclopentanyl)-carbonyl)-(R)-1,1-diamino-ethane, IV (Asp-(R)-gAla-TMCP), and aspartyl-D -valine-(R)-α-methoxymethylbenzyl amide, V (Asp-D -Val-(R)-α-methoxymethylbenzylamide). With the exception of the sodium salt II , all compounds are sweet-tasting, showing in some cases considerable potency enhancement with respect to sucrose. The results of this study confirm the earlier model that an ‘L-shape’ molecular array is essential for eliciting sweet taste for dipeptide-like ligands. In addition, it was established that (i) substitution of the N-terminal group does not inhibit sweet taste, if its zwitterionic character is maintained; (ii) a hydrophobic group located between the stem and the base of the L-shape could be responsible for sweetness potency enhancement, as found in I, III and IV ; in fact, the extraordinary potency of the N-alkylated analogue I would support a model with an additional hydrophobic binding domain above the base of the ‘L’; (iii) removal of the methyl ester at the C-terminus of compound I with the salt formation gives rise to the tasteless compound II ; (iv) for the first time all possible side-chain conformers (g⁻,g⁺andt) for the N-substituted aspartyl residue were observed; and (v) a retro-inverso modification, incorporated at position 2 of the dipeptide chain, confers greater flexibility to the molecule, as demonstrated by the contemporary presence of six conformationally distinct independent molecules in the unit cell and yet sweet taste properties are maintained, as found in IV . © 1998 European Peptide Society and John Wiley & Sons, Ltd.