世居及移居高原青少年最大有氧能力的特征

本文报道海拔３４１７ｍ和４２８０ｍ地区世居藏族和移居汉族青少年运动状态下心肺功能的对比研究。结果显示：３４１７ｍ和４２８０ｍ世居藏族的最大氧耗量、无氧阈值及最大心输出量都明显大于汉族,血氧饱和度（Ｓａｏ２）随运动负荷的增加而降低。海拔３４１７ｍ藏、汉族的△Ｓａｏ２分别为７．４６％和１０．０３％,４２８０ｍ处为８．５７％和１３．７５％,最大心率随海拔升高而下降。研究提示,藏族青少年有较高的最大有氧能力,反映了他们对低氧环境的适应优势。     

The phylogeny of the ant tribe Formicini (Hymenoptera: Formicidae) with the description of a new genus

Abstract. The holarctic ant tribe Formicini is revised, the new genus Bajcaridris described, and possible phylogenetic relationships are discussed. The subgenus Iberoformica is synonymized with Formica. A synopsis, diagnosis and keys to the genera are provided.     

Peptide analogues of the VanS catalytic center inhibit VanR binding to its cognate promoter

The dodecamer peptide SLCHDSVIGWEC, named E12, was selected from a combinatorial peptide library on the basis of its ability to bind to VanR, the two-component signal transduction response regulator which controls expression of vancomycin resistance in Enterococcus faecium. The binding of E12 was localized to the N-terminal, regulatory domain of VanR which contains Asp-55, the residue which accepts the phosphoryl group from His-164 in the activated VanS sensor kinase. E12, along with a related sequence SLAHDSIIGYLS, named E12.1, was found to inhibit the binding of VanR approximately P to a DNA segment which corresponds to its cognate promoter PvanH. With a single gap, both E12 and E12.1 could be aligned with the octadecamer sequence YLAHDIKTPLTSIIGYLS, comprising Tyr-161 through Ser-178, of the catalytic center dimerization domain of VanS, a sequence with which VanR also normally interacts. Alanine substitution analysis of E12.1 identified six amino acids as indispensable for its ability to inhibit VanR approximately P-PvanH DNA complex formation. A similar analysis of the corresponding amino acids in VanS showed a parallel dependence except for the substitutions Leu-162 --> Ala and Gly-175 --> Ala which interfered with the ability of E12.1 to compete with protein-DNA complex formation, but did not inhibit the ability of VanS to bind VanR. Our findings support a model in which E12 mimics the VanS phosphorylatable sequence with which the regulatory domain of VanR interacts, and thus functions as a "minimalist" analogue of VanS. Our results also indicate the usefulness of phage-displayed peptides as a general tool for mimicking the interacting faces of interacting proteins.     

The Aspartate-Less Receiver (ALR) Domains: Distribution,Structure and Function

Two-component signaling systems are ubiquitous in bacteria, Archaea and plants and play important roles in sensing and responding to environmental stimuli. To propagate a signaling response the typical system employs a sensory histidine kinase that phosphorylates a Receiver (REC) domain on a conserved aspartate (Asp) residue. Although it is known that some REC domains are missing this Asp residue, it remains unclear as to how many of these divergent REC domains exist, what their functional roles are and how they are regulated in the absence of the conserved Asp. Here we have compiled all deposited REC domains missing their phosphorylatable Asp residue, renamed here as the Aspartate-Less Receiver (ALR) domains. Our data show that ALRs are surprisingly common and are enriched for when attached to more rare effector outputs. Analysis of our informatics and the available ALR atomic structures, combined with structural, biochemical and genetic data of the ALR archetype RitR from Streptococcus pneumoniae presented here suggest that ALRs have reorganized their active pockets to instead take on a constitutive regulatory role or accommodate input signals other than Asp phosphorylation, while largely retaining the canonical post-phosphorylation mechanisms and dimeric interface. This work defines ALRs as an atypical REC subclass and provides insights into shared mechanisms of activation between ALR and REC domains.     

Differential assay for high-throughput screening of antibacterial compounds

The previously described Bacillus subtilis reporter strain BAU-102 is capable of detecting cell wall synthesis inhibitors that act at all stages of the cell wall synthesis pathway. In addition, this strain is capable of detecting compounds with hydrophobic/surfactant activity and alternative mechanisms of cell wall disruption. BAU-102 sequesters preformed beta-gal in the periplasm, suggesting leakage of beta-gal as the means by which this assay detects compound activities. A model is proposed according to which beta-gal release by BAU-102 reflects activation of pathways leading to autolysis. The authors also report a simplified high-throughput assay using BAU-102 combined with the fluorogenic substrate N-methylumbelliferyl-beta-D-galactoside as a single reagent. Cell wall inhibitors release beta-gal consistently only after 60 min of incubation, whereas compounds with surfactant activity show an almost immediate release. A high-throughput screen of a 480-compound library of known bioactives yielded 8 compounds that cause beta-gal release. These results validate the BAU-102 assay as an effective tool in antimicrobial drug discovery.     

Sex differences in the rate of fatigue development and recovery

Background

Many musculoskeltal injuries in the workplace have been attributed to the repetitive loading of muscle and soft tissues. It is not disputed that muscular fatigue is a risk factor for musculoskeltal injury, however the disparity between gender with respect to muscular fatigability and rate of recovery is not well understood. Current health and safety guidelines do not account for sex differences in fatiguability and may be predisposing one gender to greater risk. The purpose of this study was to quantify the sex differences in fatigue development and recovery rate of lower and upper body musculature after repeated bouts of sustained isometric contractions.

Methods

Twenty-seven healthy males (n = 12) and females (n = 15) underwent bilateral localized fatigue of either the knee extensors (male: n = 8; female: n = 8), elbow flexors (male: n = 8; female: n = 10), or both muscle groups. The fatigue protocol consisted of ten 30-second sub-maximal isometric contractions. The changes in maximum voluntary contraction (MVC), electrically evoked twitches, and motor unit activation (MUA) were assessed along with the ability to control the sustained contractions (SLP) during the fatigue protocol using a mixed four-factor repeated measures ANOVA (gender × side × muscle × time) design with significance set at p < 0.05.

Results

There was a significant loss of MVC, MUA, and evoked twitch amplitude from pre- to post-fatigue in both the arms and legs. Males had greater relative loss of isometric force, a higher rate of fatigue development, and were less capable of maintaining the fatiguing contractions in the legs when compared to the females.

Conclusion

The nature of the induced fatigue was a combination of central and peripheral fatigue that did not fully recover over a 45-minute period. The results appear to reflect sex differences that are peripheral, and partially support the muscle mass hypothesis for explaining differences in muscular fatigue.

     

Solution structure of a cyanobacterial phytochrome GAF domain in the red-light-absorbing ground state

The unique photochromic absorption behavior of phytochromes (Phys) depends on numerous reversible interactions between the bilin chromophore and the associated polypeptide. To help define these dynamic interactions, we determined by NMR spectroscopy the first solution structure of the chromophore-binding cGMP phosphodiesterase/adenylcyclase/FhlA (GAF) domain from a cyanobacterial Phy assembled with phycocyanobilin (PCB). The three-dimensional NMR structure of Synechococcus OS-B′ cyanobacterial Phy 1 in the red-light-absorbing state of Phy (Pr) revealed that PCB is bound to Cys138 of the GAF domain via the A-ring ethylidene side chain and is buried within the GAF domain in a ZZZsyn,syn,anti configuration. The D ring of the chromophore sits within a hydrophobic pocket and is tilted by approximately 80° relative to the B/C rings by contacts with Lys52 and His169. The solution structure revealed remarkable flexibility for PCB and several adjacent amino acids, indicating that the Pr chromophore has more freedom in the binding pocket than anticipated. The propionic acid side chains of rings B and C and Arg101 and Arg133 nearby are especially mobile and can assume several distinct and energetically favorable conformations. Mutagenic studies on these arginines, which are conserved within the Phy superfamily, revealed that they have opposing roles, with Arg101 and Arg133 helping stabilize and destabilize the far-red-light-absorbing state of Phy (Pfr), respectively. Given the fact that the Synechococcus OS-B′ GAF domain can, by itself, complete the Pr → Pfr photocycle, it should now be possible to determine the solution structure of the Pfr chromophore and surrounding pocket using this Pr structure as a framework.     

Cyanochromes Are Blue/Green Light Photoreversible Photoreceptors Defined by a Stable Double Cysteine Linkage to a Phycoviolobilin-type Chromophore

Phytochromes are a collection of bilin-containing photoreceptors that regulate a diverse array of processes in microorganisms and plants through photoconversion between two stable states, a red light-absorbing Pr form, and a far red light-absorbing Pfr form. Recently, a novel set of phytochrome-like chromoproteins was discovered in cyanobacteria, designated here as cyanochromes, that instead photoconvert between stable blue and green light-absorbing forms Pb and Pg, respectively. Here, we show that the distinctive absorption properties of cyanochromes are facilitated through the binding of phycocyanobilin via two stable cysteine-based thioether linkages within the cGMP phosphodiesterase/adenyl cyclase/FhlA domain. Absorption, resonance Raman and infrared spectroscopy, and molecular modeling of the Te-PixJ GAF (cGMP phosphodiesterase/adenyl cyclase/FhlA) domain assembled with phycocyanobilin are consistent with attachments to the C3¹ carbon of the ethylidene side chain and the C4 or C5 carbons in the A–B methine bridge to generate a double thioether-linked phycoviolobilin-type chromophore. These spectroscopic methods combined with NMR data show that the bilin is fully protonated in the Pb and Pg states and that numerous conformation changes occur during Pb → Pg photoconversion. Also identified were a number of photochromically inactive mutants with strong yellow or red fluorescence that may be useful for fluorescence-based cell biological assays. Phylogenetic analyses detected cyanochromes capable of different signaling outputs in a wide range of cyanobacterial species. One unusual case is the Synechocystis cyanochrome Etr1 that also binds ethylene, suggesting that it works as a hybrid receptor to simultaneously integrate light and hormone signals.Phytochromes (Phys)³ comprise a large and diverse superfamily of photoreceptors that regulate a wide range of physiological responses in plants, fungi, bacteria, and cyanobacteria (1–3). They are unique among photoreceptors by being able to photoconvert between two stable states, a red light-absorbing Pr form that is typically the dark-adapted and biologically inactive conformer and a far-red light-absorbing Pfr form that requires light for its production and is typically the biologically active conformer. By interconverting between Pr and Pfr, Phys act as light-regulated switches in controlling processes ranging from phototaxis and pigmentation in bacteria to seed germination, photomorphogenesis, and flowering time in higher plants.Light absorption by Phys is directed by a bilin (or linear tetrapyrrole) chromophore produced by the oxidative cleavage of heme. Although bacterial and fungal Phys use the immediate cleavage product biliverdin (BV), cyanobacterial and higher plant Phys use phycocyanobilin (PCB) and phytochromobilin, respectively, produced by enzymatic reduction of BV (1, 2). The bilin is then covalently bound autocatalytically to the photosensory unit of the apoprotein, which typically contains a sequence of Per/Arndt/Sim (PAS), cGMP phosphodiesterase/adenyl cyclase/FhlA (GAF), and Phy-associated (PHY) domains. Intimate contact between the bilin and surrounding protein residues then generates the unique photochromic properties of Phys. Recent three-dimensional structures of the Pr form of several bacterial Phys (BphPs) and two cyanobacterial Phys (Cphs) have shown that the bilin is deeply buried within the GAF domain in a ZZZssa configuration and that the connection between the GAF and PAS domains is stabilized by a rare figure-of-eight knot involving the region upstream of the PAS domain being lassoed by a conserved loop within the GAF domain (4–9). Although the structure of Pfr remains unsolved, various physicochemical studies have proposed that photoconversion involves a rotation of one of the three methine bridges between the pyrrole rings (1, 10–14). This rotation then induces much slower thermally driven movements of the protein to initiate signal transduction.In microorganisms, Pfr can activate a variety of signaling systems using output motifs directly appended to the C-terminal end of the photosensory region. The most prevalent are histidine kinase domains that then begin specific two-component phosphorelays (3, 15, 16). Although the output of plant Phys remains unclear, the presence of a C-terminal HK-related domain suggests that they also work as light-regulated protein kinases (17).In addition to the canonical Phys, it has become apparent through phylogenetic and biochemical studies that a heterogeneous collection of Phy-like photoreceptors exists (e.g. Refs. 3 and 18). These include Phys that prefer Pfr as the dark-adapted state (7, 19, 20), Phys that photoconvert from Pr to shorter wavelength-absorbing “near red” or Pnr forms (6, 21), and Phy-like photoreceptors that bind bilins but instead photoconvert between forms with maximal absorption other than red and far-red light (22–25). Often these Phy-like sequences are missing key residues or domains common among canonical Phys, suggesting that they employ novel bilins as chromophores, bind the bilin in different architectures, and/or use distinct photochemistries.One subclass of novel Phy-like photoreceptors present in a number of cyanobacteria, which we have designated cyanochromes (or Cycs) to better distinguish them from Cphs, is exemplified by Synechocystis sp. PCC6803 (Syn) PixJ (or TaxD1, locus sll0041) and its relatives. Syn-PixJ was discovered based on its involvement in blue light-mediated phototaxis in this mesophilic cyanobacterium (26, 27) with its close homolog Te-PixJ (locus tll0569) then found in the thermophilic cyanobacterium Thermosynechococcus elongatus BP-1 by sequence similarity (28). Like Cphs, the cyanochromes tested thus far covalently bind PCB but then generate photoreceptors that convert between blue and green light-absorbing forms designated Pb and Pg, respectively (22, 24, 29). Subsequent studies proposed that PCB is converted to phycoviolobilin (PVB) upon attachment to the apoprotein (30). PVB differs from PCB by having a methylene instead of a methine bridge between the A and B pyrrole rings, which blue-shifts the absorption of the chromophore by shortening the π-conjugation system. Phototransformation of Pb to Pg could then occur by a mechanism similar to Phys.How Te-PixJ and related cyanochromes bind PCB to generate more blue-shifted PVB-type chromophores remains unclear. Like Cphs, two cyanochromes examples link PCB via a thioether linkage between a cysteine in the Cyc-GAF domain and the C3¹ carbon of the ethylidene side chain of ring A (24, 28). Additionally, loss of the C4C5 double bond is necessary to generate PVB. One model by Ishizuka et al. (30) from studies with Te-PixJ proposed that the double bond moves from the C4-C5 position to the C2-C3 position by an autoisomerase activity intrinsic to the GAF domain. A more recent model by Rockwell et al. (24) using another Syn-PixJ relative in T. elongatus, Tlr0924, invoked the possibility of a second cysteine that also participates in PCB ligation. This cysteine was proposed to bind the bilin at the C10 position via a reversible thioether linkage. In the dark-adapted Pb state, the second linkage would then be formed to generate a rubin-like chromophore attached to the bridge between the B and C pyrrole rings. This bond would then break upon photoconversion to generate the more π-conjugated green light-absorbing photoproduct Pg.In this report, we employed a number of physicochemical approaches to help resolve the unique chromophore architecture and photochemical properties of cyanochromes, using Te-PixJ as the example. By independently mutagenizing the cysteine that binds the A ring ethylidene (Cys-522 (22)) and that proposed by Rockwell et al. (24) to reversibly bind the bilin at a second site (Cys-494), we demonstrate that both residues form light-stable covalent adducts with a PVB-type chromophore. In addition, we employed various spectroscopic methods to show that the bound PVB is fully protonated as both Pb and Pg, that only one pyrrole ring is active during photoconversion, and that the polypeptide may undergo extensive remodeling as Pb converts to Pg. We identified a set of conserved amino acids in Te-PixJ important for cyanochrome photochemistry, including several that when substituted generate yellow or red fluorescent chromoproteins potentially useful for cell biological applications. Phylogenetic analyses show that cyanochromes are widespread among cyanobacteria with their closest relatives being members of the red/far-red light-absorbing Phy subfamily defined by the absence of the N-terminal PAS domain (31).