The Power of Two-Dimensional Dwell-Time Analysis for Model Discrimination, Temporal Resolution, Multichannel Analysis and Level Detection

Two-dimensional (2D) dwell-time analysis of time series of single-channel patch-clamp current was improved by employing a Hinkley detector for jump detection, introducing a genetic fit algorithm, replacing maximum likelihood by a least square criterion, averaging over a field of 9 or 25 bins in the 2D plane and normalizing per measuring time, not per events. Using simulated time series for the generation of the “theoretical” 2D histograms from assumed Markov models enabled the incorporation of the measured filter response and noise. The effects of these improvements were tested with respect to the temporal resolution, accuracy of the determination of the rate constants of the Markov model, sensitivity to noise and requirement of open time and length of the time series. The 2D fit was better than the classical hidden Markov model (HMM) fit in all tested fields. The temporal resolution of the two most efficient algorithms, the 2D fit and the subsequent HMM/beta fit, enabled the determination of rate constants 10 times faster than the corner frequency of the low-pass filter. The 2D fit was much less sensitive to noise. The requirement of computing time is a problem of the 2D fit (100 times that of the HMM fit) but can now be handled by personal computers. The studies revealed a fringe benefit of 2D analysis: it can reveal the “true” single-channel current when the filter has reduced the apparent current level by averaging over undetected fast gating.     

Marker-Dependent Recombination in T4 Bacteriophage. III. Structural Prerequisites for Marker Discrimination

Distance- as well as marker-dependence of genetic recombination of bacteriophage T4 was studied in crosses between rIIB mutants with known base sequences. The notion of a "basic recombination," which is the recombination within distances shorter than hybrid DNA length in the absence of mismatch repair and any marker effects, was substantiated. The basic recombination frequency per base pair can serve as an objective parameter (natural constant) of general recombination reflecting its intensity. Comparative studies of the recombination properties of rIIB mutants with various sequence changes in the mutated sites showed that the main factor determining the probability of mismatch repair in recombination heteroduplexes is the length of a continuous heterologous region. A run of A:T pairs immediately adjoining the mismatch appears to stimulate its repair. In the case of mismatches with DNA strands of unequal length, formed by frameshift mutations, the repair is asymmetric, the longer strand (bulge) being preferentially removed. A pathway for mismatch repair including sequential action of endonuclease VII (gp49)----3'----5' exonuclease (gp43)----DNA polymerase (gp43)----DNA ligase (gp30) was proposed. A possible identity of the recombinational mismatch repair mechanism to that operating to produce mutations via sequence conversion is discussed.     

Structural Basis for Antibody Discrimination between Two Hormones That Recognize the Parathyroid Hormone Receptor

Parathyroid hormone-related protein (PTHrP) plays a vital role in the embryonic development of the skeleton and other tissues. When it is produced in excess by cancers it can cause hypercalcemia, and its local production by breast cancer cells has been implicated in the pathogenesis of bone metastasis formation in that disease. Antibodies have been developed that neutralize the action of PTHrP through its receptor, parathyroid hormone receptor 1, without influencing parathyroid hormone action through the same receptor. Such neutralizing antibodies against PTHrP are therapeutically effective in animal models of the humoral hypercalcemia of malignancy and of bone metastasis formation. We have determined the crystal structure of the complex between PTHrP (residues 1–108) and a neutralizing monoclonal anti-PTHrP antibody that reveals the only point of contact is an α-helical structure extending from residues 14–29. Another striking feature is that the same residues that interact with the antibody also interact with parathyroid hormone receptor 1, showing that the antibody and the receptor binding site on the hormone closely overlap. The structure explains how the antibody discriminates between the two hormones and provides information that could be used in the development of novel agonists and antagonists of their common receptor.The discovery of parathyroid hormone (PTH)6 -related protein (PTHrP) as the cause of hypercalcemia in many patients with cancer provided new insights into the pathogenesis of the skeletal complications of malignancy (1). It revealed PTHrP as a previously unrecognized hormone, related in evolution to the calcium-regulating PTH, but important in the pathogenesis of the humoral hypercalcemia of malignancy, a syndrome in which hypercalcemia occurs without evident bone metastases. Whereas PTH consists of 84 amino acids, human PTHrP has three alternative splice products of 139, 141, and 173 residues. Apart from 8 of the first 13 residues of PTH and PTHrP being identical, there is no significant identity between these peptides (2). PTHrP actively promotes bone resorption, doing so in a manner identical to that of PTH by acting upon the receptor (PTH1R) it shares with PTH. The PTH1R is located on cells of the osteoblast lineage, which program the formation and activation of osteoclasts, and on cells of the kidney tubule, through which both PTHrP and PTH promote cyclic AMP and phosphorus excretion but reduce calcium excretion. Other actions of PTHrP that reflect those of PTH include the ability to relax vascular and other smooth muscle. This response may reflect a physiological function of PTHrP rather than of PTH and is consistent with PTHrP production and local action on smooth muscles at various sites (3).The first 34 amino acids of each hormone contain the full biological activities of both PTH and of PTHrP to activate the PTH1R (4). The sequences of PTHrP and PTH between residues 14 and 34 are interesting in that, although they are not homologous, nevertheless they appear to be critical for binding of each to the seven transmembrane G protein-coupled receptor, PTH1R (4). Within the first 34 amino acids of PTH and PTHrP two functional regions have been revealed based on structural and cross-linking studies (5–8). These studies have indicated that the C-terminal half of the first 34 residues of each hormone comprises the high affinity binding domain, interacting with the N-terminal portion of the extracellular domain of the receptor. The N-terminal half of each hormone activates the receptor through contact points on the extracellular loops and juxtamembrane regions (9).Despite their equal ability to activate through the PTH1R, it was clear from the earliest work, even with antibodies against peptides within the first 14 residues of PTHrP, that highly specific antibodies could be generated that discriminate between PTH and PTHrP (10). Likewise, polyclonal antibodies against PTHrP-(1–34) that neutralized its effects completely in vitro in promotion of cyclic AMP production in response to PTHrP without any detectable neutralizing effect on PTH were used to prevent and to treat hypercalcemia in nude mice bearing xenografts of PTHrP-secreting human cancers (11, 12). Similar results were obtained with a neutralizing mouse monoclonal antibody against PTHrP (13). Subsequently, after the finding that breast cancer metastases to bone were enriched in PTHrP production (14), Guise and Mundy (15) used an experimental model in nude mice in which human breast cancer cells grow as lytic deposits in bone after intracardiac injection and showed that PTHrP production by the cancers contributed to the process of tumor establishment and growth in bone by promoting osteoclast formation and bone resorption. Furthermore, the tumor establishment and growth in bone could be prevented by treating the mice with a monoclonal antibody against PTHrP (16) or with a bisphosphonate (17) to inhibit bone resorption.The efficacy of anti-PTHrP antibodies in treating both humoral-mediated hypercalcemia in cancer and bone metastasis formation and growth in mouse models raises the prospect of humanized forms of these antibodies being used as therapeutic agents in these diseases in human subjects, and preclinical data have been obtained in support of that (18, 19). With that in mind, the present project was undertaken in which we have made use of a monoclonal antibody prepared against human PTHrP (residues 1–34), which neutralizes the actions of PTHrP through PTH1R without any action against PTH. The antibody has been complexed with recombinant human PTHrP (residues 1–108) to generate crystals that have been used to analyze the three-dimensional structure with the aim of discovering the structural basis of neutralization of PTHrP action by the antibody.     

动物辨别亲属的机制

亲缘识别在动物界中是很普遍的现象。不同的动物有不同的手段识别亲属。亲缘辨别的机制主要有空间识别机制,熟悉机制,表形匹配和识别等位基因4种机制,有的动物只有一种机制起作用,有的则是两种机制同时共存或不同时期不同机制。此外,还概述了亲缘辨别的一些功能。     

结合叶绿体和核DNA序列变异可提高云杉属物种的分子鉴别率

松科(Pinaceae)云杉属(Picea)植物是北半球广泛分布的重要森林树种,由于频繁的种间杂交渐渗及近期的辐射分化导致种间形态趋同,传统的形态学方法很难准确鉴定该属物种.近期兴起和发展的DNA条形码技术为云杉属物种的划分和鉴定提供了可参考的方法.在云杉属青藏高原种质资源收集过程完成后,选取5个叶绿体DNA片段(matK,rbcL,trnH-psbA,trnL-trnF和trnS-trnG)以及3个核DNA片段(4CL,Sb29和GI),利用PWG-distance和Tree-Building两种方法对青藏高原以及中国其他地区分布的19个云杉属物种83个个体进行了物种鉴别分辨率的评价.研究结果显示单个的叶绿体DNA片段(10.5％ ～26.3％)和核DNA片段(15.8％ ～26.3％)对云杉属物种鉴别的分辨率较低,组合的叶绿体DNA片段的分辨能力(15.8％ ～42.1％)虽然高于单个DNA片段,但分辨率最高的trnH-psbA+trnS-trnG和trnS-trnG+trn L-trnF两个组合也只能达到42.1％;组合的核DNA片段(26.3％ ～36.8％)一样对云杉属物种鉴别存在困难.但是叶绿体DNA片段和核DNA片段的组合可以明显提高对云杉属物种鉴定的分辨率,尤其是trnS-trnG+trn L-trnF+4Cl的组合片段,其分辨率可达到57.9％.因此在将来利用DNA条形码鉴别物种时,在常规DNA条形码片段不起作用的情况下,可采用这种叶绿体DNA片段和核DNA片段组合的方法来鉴定和区分植物物种.     

Power Law Distribution Defines Structural Disorder as a Structural Element Directly Linked with Function

Although intrinsically disordered proteins are prevalent and functionally important, it has never been asked whether structural disorder should be considered as a separate structural category on its own or merely as a lack of secondary and/or tertiary structure. We address this issue by showing that its length distribution in the human proteome follows a power law, with many short regions but also a significant incidence of very long disordered regions. This behavior is in sharp contrast with that of conventional secondary structural elements and is highly reminiscent of the distribution of tertiary structural units in proteins. We interpret this finding by the direct functional involvement of disorder, which distinguishes it from secondary structural elements and endows it with tertiary structural attributes.     

终止子的概念与结构之辨析

指出一些生物化学,分子生物学及遗传学方面高等院校教材和著作中,较为普遍存在的关于转录终止子概念与结构的描述和解释方面的错误。     

The Effect of Self-Efficacy on Visual Discrimination Sensitivity

Can subjective belief about one''s own perceptual competence change one''s perception? To address this question, we investigated the influence of self-efficacy on sensory discrimination in two low-level visual tasks: contrast and orientation discrimination. We utilised a pre-post manipulation approach whereby two experimental groups (high and low self-efficacy) and a control group made objective perceptual judgments on the contrast or the orientation of the visual stimuli. High and low self-efficacy were induced by the provision of fake social-comparative performance feedback and fictional research findings. Subsequently, the post-manipulation phase was performed to assess changes in visual discrimination thresholds as a function of the self-efficacy manipulations. The results showed that the high self-efficacy group demonstrated greater improvement in visual discrimination sensitivity compared to both the low self-efficacy and control groups. These findings suggest that subjective beliefs about one''s own perceptual competence can affect low-level visual processing.     

Improving the Power of Structural Variation Detection by Augmenting the Reference

The uses of the Genome Reference Consortium’s human reference sequence can be roughly categorized into three related but distinct categories: as a representative species genome, as a coordinate system for identifying variants, and as an alignment reference for variation detection algorithms. However, the use of this reference sequence as simultaneously a representative species genome and as an alignment reference leads to unnecessary artifacts for structural variation detection algorithms and limits their accuracy. We show how decoupling these two references and developing a separate alignment reference can significantly improve the accuracy of structural variation detection, lead to improved genotyping of disease related genes, and decrease the cost of studying polymorphism in a population.     

The Role of Imitation in Discrimination of Sound Series

1. Rapid sequences of vowels are discriminated considerably better than rapid sequences of tones.     

Structural Evidence that Peroxiredoxin Catalytic Power Is Based on Transition-State Stabilization

Peroxiredoxins (Prxs) are important peroxidases associated with both antioxidant protection and redox signaling. They use a conserved Cys residue to reduce peroxide substrates. The Prxs have a remarkably high catalytic efficiency that makes them a dominant player in cell-wide peroxide reduction, but the origins of their high activity have been mysterious. We present here a novel structure of human PrxV at 1.45 Å resolution that has a dithiothreitol bound in the active site with its diol moiety mimicking the two oxygens of a peroxide substrate. This suggests diols and similar di-oxygen compounds as a novel class of competitive inhibitors for the Prxs. Common features of this and other structures containing peroxide, peroxide-mimicking ligands, or peroxide-mimicking water molecules reveal hydrogen bonding and steric factors that promote its high reactivity by creating an oxygen track along which the peroxide oxygens move as the reaction proceeds. Key insights include how the active-site microenvironment activates both the peroxidatic cysteine side chain and the peroxide substrate and how it is exquisitely well suited to stabilize the transition state of the in-line S_N2 substitution reaction that is peroxidation.     

The Discrimination between Magnesium and Manganese by Serum Proteins

Magnesium and manganese have proved physically and functionally interchangeable in many isolated biological systems investigated in vitro. This lack of discrimination contrasts sharply with the high biological specificity exhibited by intact mammals under a large variety of conditions. The dichotomy between intact animals and their isolated systems might be due at least partially to presence vs. absence of an intact circulation. Hence the capability of mammalian plasma to discriminate between the alkaline earth and the transition metal was investigated by means of equilibrium dialysis, exchange, ultrafiltration, ultracentrifugation, and zone electrophoresis. The states of the respective elements are thus contrasted as follows: (a) Magnesium is partially bound, manganese totally. (b) Magnesium is nonselectively bound by serum proteins, manganese selectively by a β₁-globulin. (c) Under conditions approaching physiological, the two metals do not interchange. This is interpreted as indicating that the plasma proteins contribute to biological specificity by discriminating between a trace metal and a macronutrient.     

Haptic Discrimination of Distance

While quite some research has focussed on the accuracy of haptic perception of distance, information on the precision of haptic perception of distance is still scarce, particularly regarding distances perceived by making arm movements. In this study, eight conditions were measured to answer four main questions, which are: what is the influence of reference distance, movement axis, perceptual mode (active or passive) and stimulus type on the precision of this kind of distance perception? A discrimination experiment was performed with twelve participants. The participants were presented with two distances, using either a haptic device or a real stimulus. Participants compared the distances by moving their hand from a start to an end position. They were then asked to judge which of the distances was the longer, from which the discrimination threshold was determined for each participant and condition. The precision was influenced by reference distance. No effect of movement axis was found. The precision was higher for active than for passive movements and it was a bit lower for real stimuli than for rendered stimuli, but it was not affected by adding cutaneous information. Overall, the Weber fraction for the active perception of a distance of 25 or 35 cm was about 11% for all cardinal axes. The recorded position data suggest that participants, in order to be able to judge which distance was the longer, tried to produce similar speed profiles in both movements. This knowledge could be useful in the design of haptic devices.     

Ribosomal Discrimination of tRNAs

Mutations in two proteins of the 30S ribosomal subunit indicate that the ribosome provides a recognition screen for tRNAs before, or simultaneous with, their interaction with mRNA.     

The Structural Organization of Oligonucleosomes

Abstract

We have used electric birefringence to study the structure of oligonucleosomes and to show the influence of histone H1 depletion on their conformation in solution. Measurements are made at low ionic strength on monodisperse samples containing up to 8 nucleosomes. For each oligomer, having H1 or not, the analysis of both relaxation and orientation times gives information about the particle's orientation mechanism through the ratio r of permanent over induced dipole terms. For native oligomers, the data confirm the previous finding of a discontinuity in hydrodynamic behavior between pentamer and heptamer: the rotational times are multiplied by 10 and r increases from 0.2 to 0.7 showing the appearance of a non-negligible contribution of a permanent dipole to the orientation mechanism. We suggest a model for the hexanucleosome at low ionic strength and discuss its implications for the higher-order structure of chromatin.

The treatment for H1 depletion abolishes the transitions in electro-optical properties: the value of r remains constant, r=0.15, and both rotational times increase progressively with the number of nucleosomes in the chain. That reflects an important unfolding of oligonucleosomal structure which we attributed to the unwinding of DNA tails and internucleosomal segments. The disc planes of nucleosomes become closely parallel to the nucleosomal chain axis.     

The Mexican Outsiders: A Community History of Marginalization and Discrimination

The Mexican Outsiders:. Community History of Marginalization and Discrimination. Martha Menchaca. Austin: University of Texas Press, 1995. 250 pp.