A review of recent advances in the study of early angiosperms from northeastern China

The last 10 years (1998–2007) were very productive and important in the study of early angiosperms in northeastern China. The new discoveries of the earliest well-documented records of angiosperms such as Archaefructus, as well as Hyrcantha decussata (= Sinocarpus decussatus), provided fresh knowledge for better understanding the primitive characters of the ancient angiosperms and also their aquatic (or wet) habitat and their herbaceous nature. Some new approaches such as the combination of molecular and morphological characters joined together to place Archaefructus in the angiosperm phylogenetic framework. These fossils demonstrate that we should expect more ancient angiosperms to be found in the pre-Cretaceous which will continue to add important new understanding to the nature of the origin and evolution of the angiosperms.     

Paleozoic Protein Fossils Illuminate the Evolution of Vertebrate Genomes and Transposable Elements

Genomes hold a treasure trove of protein fossils: Fragments of formerly protein-coding DNA, which mainly come from transposable elements (TEs) or host genes. These fossils reveal ancient evolution of TEs and genomes, and many fossils have been exapted to perform diverse functions important for the host’s fitness. However, old and highly degraded fossils are hard to identify, standard methods (e.g. BLAST) are not optimized for this task, and few Paleozoic protein fossils have been found. Here, a recently optimized method is used to find protein fossils in vertebrate genomes. It finds Paleozoic fossils predating the amphibian/amniote divergence from most major TE categories, including virus-related Polinton and Gypsy elements. It finds 10 fossils in the human genome (eight from TEs and two from host genes) that predate the last common ancestor of all jawed vertebrates, probably from the Ordovician period. It also finds types of transposon and retrotransposon not found in human before. These fossils have extreme sequence conservation, indicating exaptation: some have evidence of gene-regulatory function, and they tend to lie nearest to developmental genes. Some ancient fossils suggest “genome tectonics,” where two fragments of one TE have drifted apart by up to megabases, possibly explaining gene deserts and large introns. This paints a picture of great TE diversity in our aquatic ancestors, with patchy TE inheritance by later vertebrates, producing new genes and regulatory elements on the way. Host-gene fossils too have contributed anciently conserved DNA segments. This paves the way to further studies of ancient protein fossils.     

Paleoproteomics of Mesozoic Dinosaurs and Other Mesozoic Fossils

Molecular studies have contributed greatly to our understanding of evolutionary processes that act upon virtually every aspect of living organisms. However, these studies are limited with regard to extinct organisms, particularly those from the Mesozoic because fossils pose unique challenges to molecular workflows, and because prevailing wisdom suggests no endogenous molecular components can persist into deep time. Here, the power and potential of a molecular approach to Mesozoic fossils is discussed. Molecular methods that have been applied to Mesozoic fossils—including iconic, non‐avian dinosaurs— and the challenges inherent in such analyses, are compared and evaluated. Taphonomic processes resulting in the transition of living organisms from the biosphere into the fossil record are reviewed, and the possible effects of taphonomic alteration on downstream analyses that can be problematic for very old material (e.g., molecular modifications, limitations of on comparative databases) are addressed. Molecular studies applied to ancient remains are placed in historical context, and past and current studies are evaluated with respect to producing phylogenetically and/or evolutionarily significant data. Finally, some criteria for assessing the presence of endogenous biomolecules in very ancient fossil remains are suggested as a starting framework for such studies.     

古蛋白质分析在东亚古人类演化中的应用前景

东亚古人类演化是学术界关注的热点科学问题,国内外学者对此进行了多学科的相关研究,取得了很多重要进展,但仍然存在许多尚未解决的问题。古蛋白质分析近年来成为古生物演化领域的又一个前沿和热点方向,取得了一系列重要突破。较之古DNA,古蛋白质的保存优势使其可以在时间上和地域上突破古DNA的限制,在古人类演化领域大有可为。东亚古人类化石丰富且时段大致连续,但更新世或更早时期的分子证据非常缺乏。本文从古蛋白质分析的发展史、研究潜力、难点与挑战以及思考与展望等几方面,对古蛋白质分析在东亚古人类演化研究中的应用前景进行梳理与思考。相信随着更多分子证据的积累,古蛋白质分析可为东亚古人类的演化脉络提供更多关键性的线索,极大地促进人类演化研究。     

What Does Palaeontology Contribute to Systematics in a Molecular World?

Palaeontology provides the only direct record for morphological and genetic change through time and uniquely contributes to systematics in two ways: by providing access to denser taxon sampling than is otherwise possible and by dating divergence times. Claims that ancient DNA has survived millions of years in certain fossils suggested the possibility that palaeontology could contribute directly to molecular systematic studies. Unfortunately, none of the supposed geologically ancient DNA records stands up to detailed scrutiny and fossils therefore contribute primarily through the morphological information they preserve. Denser taxon sampling can improve the accuracy of phylogenetic estimates primarily through allowing better discrimination of homoplasy from homology. This in turn leads to more accurate hypotheses of character transformation. Denser taxon sampling also offers the opportunity for more accurate rooting, since more characters can be polarized by reference to a stem-group taxon than to an extant sister-group taxon. Missing data can be a problem for fossils, but is not crippling. Finally the temporal order of clade appearances in the fossil record can provide ancillary evidence for selecting a working phylogeny from among a number of equally most parsimonious cladograms.     

Are pollen fossils useful for calibrating relaxed molecular clock dating of phylogenies? A comparative study using Myrtaceae

The identification and application of reliable fossil calibrations represents a key component of many molecular studies of evolutionary timescales. In studies of plants, most paleontological calibrations are associated with macrofossils. However, the pollen record can also inform age calibrations if fossils matching extant pollen groups are found. Recent work has shown that pollen of the myrtle family, Myrtaceae, can be classified into a number of morphological groups that are synapomorphic with molecular groups. By assembling a data matrix of pollen morphological characters from extant and fossil Myrtaceae, we were able to measure the fit of 26 pollen fossils to a molecular phylogenetic tree using parsimony optimisation of characters. We identified eight Myrtaceidites fossils as appropriate for calibration based on the most parsimonious placements of these fossils on the tree. These fossils were used to inform age constraints in a Bayesian phylogenetic analysis of a sequence alignment comprising two sequences from the chloroplast genome (matK and ndhF) and one nuclear locus (ITS), sampled from 106 taxa representing 80 genera. Three additional analyses were calibrated by placing pollen fossils using geographic and morphological information (eight calibrations), macrofossils (five calibrations), and macrofossils and pollen fossils in combination (12 calibrations). The addition of new fossil pollen calibrations led to older crown ages than have previously been found for tribes such as Eucalypteae and Myrteae. Estimates of rate variation among lineages were affected by the choice of calibrations, suggesting that the use of multiple calibrations can improve estimates of rate heterogeneity among lineages. This study illustrates the potential of including pollen-based calibrations in molecular studies of divergence times.     

根据蛋白质的氨基酸组成实现其快速鉴定

常规进行蛋白质鉴定的方法是测定其氨基酸顺序,它需要蛋白质顺序分析仪,对蛋白质的纯度要求高,费时和花费大,与之相比,蛋白质的氨基酸组成和分子量是容易实验测定的。本文描述了一个基于蛋白质的组成和分子量进行其快速鉴定的方法。其基本出发点是,通过统计蛋白质序列数据库中每个序列的氨基酸组成和分子量,得到一个含蛋白质长度、组成和分子量的数据库,将靶蛋白质的组成等数据与该数据库进行对比,可以检出组成和分子量与之接近的蛋白质。从而对该蛋白质进行初步鉴定。在有些情况下,甚至能相当准确地确定靶蛋白质与数据库中的某个（些）蛋白质相关。根据这一原理本文设计了根据氨基酸组成检索蛋白质组成数据库的程序,通过对胰岛素原、细胞肿瘤抗原Ｐ５３和泛肽等多种蛋白质的组成分析,证实根据氨基酸组成能较好地进行蛋白质鉴定。     

以胶原蛋白为例浅述古蛋白质组学研究现状与未来*

蛋白质作为参与构建生物体的重要生物大分子,是生物功能与代谢的物质基础,同时蛋白质的序列信息又源自生物的遗传编码信息,因此是认识生物演化本质的重要研究对象。近年来,随着质谱技术的发展,获取生物化石中的古蛋白质序列信息不再遥不可及,这为在形态学与DNA序列信息缺乏的条件下对古生物的认识提供了一条新途径。胶原蛋白(Collagen)在动物骨骼中极为丰富,又因为其特殊的结构,易于在化石中保存,故已成为古蛋白质组学研究的重要对象。本文将以胶原蛋白为例,对古蛋白质组学现有的研究方法与已经取得的研究成果进行总结,并对古蛋白质组学目前面临的挑战与困难及未来研究趋势进行讨论,旨在展示古蛋白质组学的应用潜力,并探讨其在生物演化研究中的意义。     

Monocrotaline pyrrole targets proteins with and without cysteine residues in the cytosol and membranes of human pulmonary artery endothelial cells

A single injection of monocrotaline produces a pulmonary insult in rats with a phenotype similar to human primary pulmonary hypertension. Although extensively used as a model, the mechanism(s) by which this chemical insult mimics a condition with genetic and environmental links remains an enigma, although formation of protein adducts has been implicated. Monocrotaline (MCT) is non-toxic and must undergo hepatic dehydrogenation to the soft electrophile monocrotaline pyrrole as prerequisite to damaging endothelial cells lining arterioles at remote pulmonary sites. In this report we extend our earlier investigation (J. Biol. Chem. 2000, 275, 29091-29099) by examining protein adducts to lower abundance adducts, a pI range not covered before, and subcellular localization of adduct-forming proteins associated with plasma membranes. Human pulmonary artery endothelial cells were exposed to [(14)C]MCT pyrrole (MCTP) and protein targets were identified using 2-DE with IPG 4-11. Adducted proteins were identified by pI, apparent molecular weight, and PMF using MALDI-TOF MS. Results of this study show that the majority of adducts form on proteins that contain reactive thiols in a CXXC motif, such as protein disulfide isomerase A(3) (ERp57), protein disulfide isomerase (PDI), and endothelial PDI. These same proteins were the major adduct-forming proteins associated with the plasma membrane. Other proteins found to be targets were thioredoxin, galectin-1, reticulocalbin 1 and 3, cytoskeletal tropomyosin, mitochondrial ATP synthase beta-chain, annexin A2 and cofilin-1. For the first time, MCTP adducts were observed on proteins not known to contain cysteine residues. However, known reactive proteins including nucleophosmin did not form detectable adducts, potentially indicating that MCTP did not reach the interior of nucleus to the same extent as other cellular sites. These findings suggest that molecular events underlying MCTP toxicity are initiated at the plasma membrane or readily accessible subcellular regions including the cytosol and membranes of the endoplasmic reticulum and mitochondria.     

Protein Phosphorylation in Astrocytes Mediated by Protein Kinase C: Comparison with Phosphorylation by Cyclic AMP-Dependent Protein Kinase

The protein kinase C activator, phorbol 12-myristate 13-acetate (PMA), has been found recently to transform cultured astrocytes from flat, polygonal cells into stellate-shaped, process-bearing cells. Studies were conducted to determine the effect of PMA on protein phosphorylation in astrocytes and to compare this pattern of phosphorylation with that elicited by dibutyryl cyclic AMP (dbcAMP), an activator of the cyclic AMP-dependent protein kinase which also affects astrocyte morphology. Exposure to PMA increased the amount of 32P incorporation into several phosphoproteins, including two cytosolic proteins with molecular weights of 30,000 (pI 5.5 and 5.7), an acidic 80,000 molecular weight protein (pI 4.5) present in both the cytosolic and membrane fractions, and two cytoskeletal proteins with molecular weights of 60,000 (pI 5.3) and 55,000 (pI 5.6), identified as vimentin and glial fibrillary acidic protein, respectively. Effects of PMA on protein phosphorylation were not observed in cells depleted of protein kinase C. In contrast to the effect observed with PMA, treatment with dbcAMP decreased the amount of 32P incorporation into the 80,000 protein. Like PMA, treatment with dbcAMP increased the 32P incorporation into the proteins with molecular weights of 60,000, 55,000 and 30,000, although the magnitude of this effect was different. The effect of dbcAMP on protein phosphorylation was still observed in cells depleted of protein kinase C. The results suggest that PMA, via the activation of protein kinase C, can alter the phosphorylation of a number of proteins in astrocytes, and some of these same phosphoproteins are also phosphorylated by the cyclic AMP-dependent mechanisms.     

蛋白质二级结构指定

蛋白质二级结构是指蛋白质骨架结构中有规律重复的构象。由蛋白质原子坐标正确地指定蛋白质二级结构是分析蛋白质结构与功能的基础,二级结构的指定对于蛋白质分类、蛋白质功能模体的发现以及理解蛋白质折叠机制有着重要的作用。并且蛋白质二级结构信息广泛应用到蛋白质分子可视化、蛋白质比对以及蛋白质结构预测中。目前有超过20种蛋白质二级结构指定方法,这些方法大体可以分为两大类:基于氢键和基于几何,不同方法指定结果之间的差异较大。由于尚没有蛋白质二级结构指定方法的综述文献,因此,本文主要介绍和总结已有蛋白质二级结构指定方法。     

基于蛋白质受体的药物分子计算机辅助设计策略

药物分子计算机辅助设计是一种在计算机或者理论上通过构建具有一定潜在药理活性的新化学实体的分子模拟方法。近十几年来,高通量组学技术的快速发展为生物和化学药物分子设计提供了良好的数据支撑和研究契机。另外,现代社会对生物制药合理性以及作用机理理解的要求越来越高,行业普遍要求药物需要有高效、无毒或者低毒以及靶向性强等特点。随着越来越多与药物靶点相关的蛋白质结构通过实验方法解析出来,基于蛋白质受体的药物分子设计方法可行性进一步提高,其方法也变得越来越重要。基于蛋白质受体的药物分子设计方法,一般是以蛋白质以及配体的三维结构出发进行分析,这让药物分子先导物的发现更加理性化。随着相关实验数据的积累以及深度学习等算法的发展,从而可以进行更加科学的药物分子设计,这在一定程度上加快了新药研发的进程,并更有利于探索相应的分子机理。本文对基于蛋白质受体的药物分子设计方法的常用策略进行系统的回顾、总结和展望。     

蛋白质间相互作用技术的研究近况

蛋白质间相互作用技术的研究近况黄翠芬叶棋浓（军事医学科学院生物工程研究所,北京１００８５０关键词：蛋白质,相互作用,技术ＲｅｃｅｎｔＡｄｖａｎｃｅｓｉｎｔｈｅＴｅｃｈｎｉｑｕｅｓｏｆＰｒｏｔｅｉｎ┐ＰｒｏｔｅｉｎＩｎｔｅｒａｃｔｉｏｎｓＨｕａｎｇＣｕ...     

Identification of related proteins with weak sequence identity using secondary structure information

Molecular modeling of proteins is confronted with the problem of finding homologous proteins, especially when few identities remain after the process of molecular evolution. Using even the most recent methods based on sequence identity detection, structural relationships are still difficult to establish with high reliability. As protein structures are more conserved than sequences, we investigated the possibility of using protein secondary structure comparison (observed or predicted structures) to discriminate between related and unrelated proteins sequences in the range of 10%-30% sequence identity. Pairwise comparison of secondary structures have been measured using the structural overlap (Sov) parameter. In this article, we show that if the secondary structures likeness is >50%, most of the pairs are structurally related. Taking into account the secondary structures of proteins that have been detected by BLAST, FASTA, or SSEARCH in the noisy region (with high E: value), we show that distantly related protein sequences (even with <20% identity) can be still identified. This strategy can be used to identify three-dimensional templates in homology modeling by finding unexpected related proteins and to select proteins for experimental investigation in a structural genomic approach, as well as for genome annotation.     

On the analysis of protein self-association by sedimentation velocity analytical ultracentrifugation

Analytical ultracentrifugation is one of the classical techniques for the study of protein interactions and protein self-association. Recent instrumental and computational developments have significantly enhanced this methodology. In this paper, new tools for the analysis of protein self-association by sedimentation velocity are developed, their statistical properties are examined, and considerations for optimal experimental design are discussed. A traditional strategy is the analysis of the isotherm of weight-average sedimentation coefficients s(w) as a function of protein concentration. From theoretical considerations, it is shown that integration of any differential sedimentation coefficient distribution c(s), ls-g(*)(s), or g(s(*)) can give a thermodynamically well-defined isotherm, as long as it provides a good model for the sedimentation profiles. To test this condition for the g(s(*)) distribution, a back-transform into the original data space is proposed. Deconvoluting diffusion in the sedimentation coefficient distribution c(s) can be advantageous to identify species that do not participate in the association. Because of the large number of scans that can be analyzed in the c(s) approach, its s(w) values are very precise and allow extension of the isotherm to very low concentrations. For all differential sedimentation coefficients, corrections are derived for the slowing of the sedimentation boundaries caused by radial dilution. As an alternative to the interpretation of the isotherm of the weight-average s value, direct global modeling of several sedimentation experiments with Lamm equation solutions was studied. For this purpose, a new software SEDPHAT is introduced, allowing the global analysis of several sedimentation velocity and equilibrium experiments. In this approach, information from the shape of the sedimentation profiles is exploited, which permits the identification of the association scheme and requires fewer experiments to precisely characterize the association. Further, under suitable conditions, fractions of incompetent material that are not part of the reversible equilibrium can be detected.     

Ancient fossil specimens of extinct species are genetically more distant to an outgroup than extant sister species are

There exists a remarkable correlation between genetic distance as measured by protein or DNA dissimilarity and time of species divergence as inferred from fossil records. This observation has provoked the molecular clock hypothesis. However, data inconsistent with the hypothesis have steadily accumulated in recent years from studies of extant organisms. Here the published DNA and protein sequences from ancient fossil specimens were examined to see if they would support the molecular clock hypothesis. The hypothesis predicts that ancient specimens cannot be genetically more distant to an outgroup than extant sister species are. Also, two distinct ancient specimens cannot be genetically more distant than their extant sister species are. The findings here do not conform to these predictions. Neanderthals are more distant to chimpanzees and gorillas than modern humans are. Dinosaurs are more distant to frogs than extant birds are. Mastodons are more distant to opossums than other placental mammals are. The genetic distance between dinosaurs and mastodons is greater than that between extant birds and mammals. Therefore, while the molecular clock hypothesis is consistent with some data from extant organisms, it has yet to find support from ancient fossils. Far more damaging to the hypothesis than data from extant organisms, which merely question the constancy of mutation rate, the study of ancient fossil organisms here challenges for the first time the fundamental premise of modern evolution theory that genetic distances had always increased with time in the past history of life on Earth.     

The Synthesis and Possible Transport of Specific Proteins by Cells Associated with Brain Capillaries

Abstract: Vinblastine causes alterations in the subcellular distribution of certain proteins synthesized by telencephalon slices. Proteins in various subcellular fractions were separated according to their molecular weight by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and radioactive proteins were determined by autofluorography. A microvascular fraction contained very high amounts of radioactivity in proteins with a molecular weight of 71,000. At least one of these proteins accumulated in the microvascular fraction when the telencephalon slices were incubated in vinblastine. At the same time these proteins became depleted in a myelinated axon fraction, microsomal fraction, and soluble/cytosol fraction. Vinblastine also affected the subcellular distribution of some proteins with a molecular weight below 27,000, but unlike the proteins of mol. wt. 71,000 none of these were synthesized at very high rates. Vinblastine did not effect the synthesis of protein in telencephalon slices, nor did it alter the subcellular fractionation of particles and organelles from slices. It is suggested that a non-neuronal vinblastine-sensitive protein translocation system is functioning within the cells of the microvascular network in telencephalon slices, and that at least one protein of 71,000 molecular weight and one protein with a molecular weight below 27,000 are transported on this system.     

Analysis and Prediction of the Metabolic Stability of Proteins Based on Their Sequential Features,Subcellular Locations and Interaction Networks

The metabolic stability is a very important idiosyncracy of proteins that is related to their global flexibility, intramolecular fluctuations, various internal dynamic processes, as well as many marvelous biological functions. Determination of protein''s metabolic stability would provide us with useful information for in-depth understanding of the dynamic action mechanisms of proteins. Although several experimental methods have been developed to measure protein''s metabolic stability, they are time-consuming and more expensive. Reported in this paper is a computational method, which is featured by (1) integrating various properties of proteins, such as biochemical and physicochemical properties, subcellular locations, network properties and protein complex property, (2) using the mRMR (Maximum Relevance & Minimum Redundancy) principle and the IFS (Incremental Feature Selection) procedure to optimize the prediction engine, and (3) being able to identify proteins among the four types: “short”, “medium”, “long”, and “extra-long” half-life spans. It was revealed through our analysis that the following seven characters played major roles in determining the stability of proteins: (1) KEGG enrichment scores of the protein and its neighbors in network, (2) subcellular locations, (3) polarity, (4) amino acids composition, (5) hydrophobicity, (6) secondary structure propensity, and (7) the number of protein complexes the protein involved. It was observed that there was an intriguing correlation between the predicted metabolic stability of some proteins and the real half-life of the drugs designed to target them. These findings might provide useful insights for designing protein-stability-relevant drugs. The computational method can also be used as a large-scale tool for annotating the metabolic stability for the avalanche of protein sequences generated in the post-genomic age.     

Molecular Insights into Mammalian End-binding Protein Heterodimerization

Microtubule plus-end tracking proteins (+TIPs) are involved in many microtubule-based processes. End binding (EB) proteins constitute a highly conserved family of +TIPs. They play a pivotal role in regulating microtubule dynamics and in the recruitment of diverse +TIPs to growing microtubule plus ends. Here we used a combination of methods to investigate the dimerization properties of the three human EB proteins EB1, EB2, and EB3. Based on Förster resonance energy transfer, we demonstrate that the C-terminal dimerization domains of EBs (EBc) can readily exchange their chains in solution. We further document that EB1c and EB3c preferentially form heterodimers, whereas EB2c does not participate significantly in the formation of heterotypic complexes. Measurements of the reaction thermodynamics and kinetics, homology modeling, and mutagenesis provide details of the molecular determinants of homo- versus heterodimer formation of EBc domains. Fluorescence spectroscopy and nuclear magnetic resonance studies in the presence of the cytoskeleton-associated protein-glycine-rich domains of either CLIP-170 or p150^glued or of a fragment derived from the adenomatous polyposis coli tumor suppressor protein show that chain exchange of EBc domains can be controlled by binding partners. Extension of these studies of the EBc domains to full-length EBs demonstrate that heterodimer formation between EB1 and EB3, but not between EB2 and the other two EBs, occurs both in vitro and in cells as revealed by live cell imaging. Together, our data provide molecular insights for rationalizing the dominant negative control by C-terminal EB domains and form a basis for understanding the functional role of heterotypic chain exchange by EBs in cells.