Recurring structural motifs in proteins with different functions

BACKGROUND: Structures that have diverged from a common ancestor often retain functional and sequence similarity, although the latter may be very reduced. Even so, the overall fold of the structure is generally highly conserved. Now however, several have been identified of proteins that have been identified that have different functions but which have converged to a similar fold. These proteins will also have low sequence identities. RESULTS: By comparing the complete structure databank against itself, using sequence and structure alignment techniques, we have been able to identify six new examples of structurally related folds that have no apparent sequence or functional similarity. These related proteins include a family of crambin-like folds and a family of ferredoxin II folds. We found that all the similarities between structures are present in small proteins and occur as motifs within the core of a larger protein. CONCLUSION: The low sequence similarity and the lack of any obvious functional relationship between proteins with similar structures suggest that the proteins have diverged from independent ancestors. The similarities may therefore be of interest for understanding the various stereochemical and physical criteria that operate to generate a favourable fold.     

植物硫转运蛋白研究进展

硫转运蛋白在植物对硫酸盐的吸收和转运中起着重要的作用。已经在拟南芥、大麦和小麦等植物中分离到了40多种硫转运蛋白基因。这些基因序列与其他种类生物的硫转运蛋白基因序列有着高度的保守性。利用CLUSTAL程序建立的系统进化树将植物硫转运蛋白划分为5个亚群。使用多种拓扑预测程序推测出不同植物硫转运蛋白的共同结构特点是均含有12个跨膜域。在柱花草和大麦中,硫转运蛋白基因表达调控包括植物体内硫水平的负调控和O—乙酰丝氨酸的正调控两种方式。对硫转运蛋白的组织定位和功能研究表明,高亲和硫转运蛋白主要定位于根部,在根系硫酸盐吸收中起重要作用。     

Means and extremes: building variability into community‐level climate change experiments

Experimental studies assessing climatic effects on ecological communities have typically applied static warming treatments. Although these studies have been informative, they have usually failed to incorporate either current or predicted future, patterns of variability. Future climates are likely to include extreme events which have greater impacts on ecological systems than changes in means alone. Here, we review the studies which have used experiments to assess impacts of temperature on marine, freshwater and terrestrial communities, and classify them into a set of ‘generations’ based on how they incorporate variability. The majority of studies have failed to incorporate extreme events. In terrestrial ecosystems in particular, experimental treatments have reduced temperature variability, when most climate models predict increased variability. Marine studies have tended to not concentrate on changes in variability, likely in part because the thermal mass of oceans will moderate variation. In freshwaters, climate change experiments have a much shorter history than in the other ecosystems, and have tended to take a relatively simple approach. We propose a new ‘generation’ of climate change experiments using down‐scaled climate models which incorporate predicted changes in climatic variability, and describe a process for generating data which can be applied as experimental climate change treatments.     

Development of a gas chromatography mass spectrometry assay for the two eosinophil chemotactic factors of anaphylaxis

Gas chromatography mass spectrometry has been used for over a decade for the determination of the amino acid sequences of fragment peptides derived from larger parent molecules. The majority of these fragments have from four to seven residues and several different methods of derivatization have been devised. Few reports have been published in which similar techniques have been used for the quantification of such peptides, but there is a growing list of small peptides which have been shown to have biological activity in their own right. This report is concerned with the development of a gas chromatographic mass spectrometric assay for the two eosinophil chemotactic peptides, Val-Gly-Ser-Glu and Ala-Gly-Ser-Glu, which appear to have a role to play in the course of the inflammatory process in skin disorders.     

Host defense mechanisms of cephalopods

Humoral and cellular mechanisms of defense have been described for cephalopods, a relatively advanced group of mollusks. Typical of other mollusks, cephalopod agglutinins are the most documented component of humoral immunity. Lectins, which have agglutinating properties, have been described and characterized from octopuses. Agglutinins from cephalopod hemolymph have also been shown to agglutinate a variety of vertebrate red blood cells, as well as potential bacterial pathogens. Hemocytes are the primary component of cellular immunity. Although the hemocyte role in phagocytosis has been extensively studied in other mollusks, the mechanisms of phagocytosis have not been described extensively for cephalopods. Cephalopod hemocytes have phagocytic capabilities and may function in encapsulation and neutralization of foreign substances; however, the effects of environmental factors and the full extent of phagocytic capabilities of cephalopod hemocytes have not been reported. Hemocytes from cephalopods have a role in wound healing and inflammation which have been reported in detail by several investigators.     

Comparing G matrices: are common principal components informative?

Common principal components (CPC) analysis is a technique for assessing whether variance-covariance matrices from different populations have similar structure. One potential application is to compare additive genetic variance-covariance matrices, G. In this article, the conditions under which G matrices are expected to have common PCs are derived for a two-locus, two-allele model and the model of constrained pleiotropy. The theory demonstrates that whether G matrices are expected to have common PCs is largely determined by whether pleiotropic effects have a modular organization. If two (or more) populations have modules and these modules have the same direction, the G matrices have a common PC, regardless of allele frequencies. In the absence of modules, common PCs exist only for very restricted combinations of allele frequencies. Together, these two results imply that, when populations are evolving, common PCs are expected only when the populations have modules in common. These results have two implications: (1) In general, G matrices will not have common PCs, and (2) when they do, these PCs indicate common modular organization. The interpretation of common PCs identified for estimates of G matrices is discussed in light of these results.     

Biomass estimation of orange roughy: a summary and evaluation of techniques for measuring stock size of a deep-water fish species in New Zealand

A number of techniques have been employed to measure biomass of orange roughy in New Zealand. Time series of stratified random trawl surveys in a number of areas over periods of 5–10 years have given relative abundance indices, which have been used in stock reduction analyses to estimate true biomass. This has given confident results for the Chatham Rise stock, and surveys have also shown strong trends in other regions. Acoustic surveys have been carried out on three grounds. They can give relative indices of orange roughy abundance in areas of fiat or medium sloping bottom, and have the potential for estimation of absolute biomass from a single survey. Egg production surveys have been employed on two grounds on the east coast of the North Island, where conditions of tight spawning aggregations and steep bottom topography have limited the success of other methods. Both daily fecundity reduction and annual egg production methods have been used. These have given estimates of true biomass from one-off surveys, although results are imprecise. Both unstandardized and standardized analyses of commercial catch-per-unit-effort data have given relative indices of abundance, which have formed an important part of stock assessment for several fisheries. No single technique used to measure the size of orange roughy stocks has proven ideal or appropriate in all New Zealand situations. All have advantages and disadvantages, depending on the characteristics of the fishing area and fish behaviour. For two areas, a combination of methods have been applied, which has given more confident results than those from a single technique.     

The neutral theory in the genomic era

A number of tests have been developed to detect positive selection at the molecular level. These tests are based on DNA polymorphism within and divergence between species. Applications of these tests have revealed a large collection of genes that have evolved under positive selection and some general insights into adaptive evolution. Recently, these tests have been applied on a genomic scale and have provided estimates of the frequency of adaptive substitutions and a critical test of the neutral theory.     

Pharmacology of azoles

Azole antifungals have different pharmacokinetic characteristics: complete oral absorption for Voriconazole, and to a lesser extent for fluconazole. The absorption of posaconazole and itraconazole increases with food intake. All of them have high tissue distribution with low plasma concentrations, especially low in the case of posaconazole and itraconazole. Posaconazole and itraconazole have high plasmatic protein binding and consequently both have a very low free fraction. Elimination of azole antifungals is through a metabolic pathway with CYP450 isoenzymes, and has a non linear pharmacokinetics with a high risk of interation with other drugs since azoles have the ability of CYP450 isoenzymes inhibition. Possibly the parameter that defines more precisely their efficacy is AUIC with an optimum value near 20, although cut-off values must be defined since some azoles may have difficulty to reach this value.     

Relating structure to thermodynamics: the crystal structures and binding affinity of eight OppA-peptide complexes.

The oligopeptide-binding protein OppA provides a useful model system for studying the physical chemistry underlying noncovalent interactions since it binds a variety of readily synthesized ligands. We have studied the binding of eight closely related tripeptides of the type Lysine-X-Lysine, where X is an abnormal amino acid, by isothermal titration calorimetry (ITC) and X-ray crystallography. The tripeptides fall into three series of ligands, which have been designed to examine the effects of small changes to the central side chain. Three ligands have a primary amine as the second side chain, two have a straight alkane chain, and three have ring systems. The results have revealed a definite preference for the binding of hydrophobic residues over the positively charged side chains, the latter binding only weakly due to unfavorable enthalpic effects. Within the series of positively charged groups, a point of lowest affinity has been identified and this is proposed to arise from unfavorable electrostatic interactions in the pocket, including the disruption of a key salt bridge. Marked entropy-enthalpy compensation is found across the series, and some of the difficulties in designing tightly binding ligands have been highlighted.     

Host PrP glycosylation: a major factor determining the outcome of prion infection

The expression of the prion protein (PrP) is essential for transmissible spongiform encephalopathy (TSE) or prion diseases to occur, but the underlying mechanism of infection remains unresolved. To address the hypothesis that glycosylation of host PrP is a major factor influencing TSE infection, we have inoculated gene-targeted transgenic mice that have restricted N-linked glycosylation of PrP with three TSE strains. We have uniquely demonstrated that mice expressing only unglycosylated PrP can sustain a TSE infection, despite altered cellular location of the host PrP. Moreover we have shown that brain material from mice infected with TSE that have only unglycosylated PrP^Sc is capable of transmitting infection to wild-type mice, demonstrating that glycosylation of PrP is not essential for establishing infection within a host or for transmitting TSE infectivity to a new host. We have further dissected the requirement of each glycosylation site and have shown that different TSE strains have dramatically different requirements for each of the glycosylation sites of host PrP, and moreover, we have shown that the host PrP has a major role in determining the glycosylation state of de novo generated PrP^Sc.     

The place of metabolism in the origin of life

Metabolism and replication are generally considered essential features of biological life. Workers in the field of the origin of life are mostly split into two groups, depending on which of these two functions is postulated to have occurred first. Because of difficulties encountered by the replication-first (or genetics-first) approach, some workers have postulated that a highly developed metabolism must have originated before replication and the formation of a genetic apparatus. However, as supporters of a replication-first approach have pointed out, and as is discussed in this article, the alternative metabolism-first approach has fundamental problems that have not been sufficiently addressed.     

Seven new bibenzyls and a dihydrochalcone from Radula variabilis

Six new bibenzyls have been isolated from the liverwort, Radula variabilis and their structures have been established by chemical and spectral evidence. In addition, a new dihydrochalcone and a new bibenzy] have been isolated as their methyl ethers. Except for two of the bibenzyls, the present compounds have a unique seven-membered heterocyclic ring and they are the first members of a new group of natural product.     

Design and synthesis of mono- and multimeric targeted radiopharmaceuticals based on novel cyclen ligands coupled to anti-MUC1 aptamers for the diagnostic imaging and targeted radiotherapy of cancer

Targeted radiopharmaceuticals offer the possibility of improved tumor imaging and radiotherapy, with reduced side effects. A variety of monoclonal antibodies and antibody fragments have previously been successfully radiolabeled and used in diagnostic imaging and targeted radiotherapy of cancer. Many such antibodies have been shown to recognize the well-characterized MUC1 tumor marker and have recently been in clinical trials. Furthermore, a number of chelators have been synthesized and are currently used as radiopharmaceuticals for imaging and therapy. We now report the synthesis of a novel, cyclen-based ligand with a sulfur-containing arm that offers increased stability of the ligand-metal complex. We have coupled this ligand with previously selected aptamers to the MUC1 tumor marker to generate a novel targeted radiopharmaceutical with improved properties. We have tested the complex against known, commercially available chelators such as MAG3 in model breast cancer systems. To improve the pharmacokinetic properties of the aptamer-based targeted radiopharmaceutical, we have generated multi-aptamer complexes around a central chelator. Such multi-aptamer complexes have increased retention of the complex in circulation, without affecting the lack of immunogenicity of the complex or altering its superior tumor penetration properties.     

Protein redesign.

It has been demonstrated that, for a number of proteins, it is possible to dramatically alter the connectivities between elements of secondary structure. Remarkably large loop insertions are tolerated and many redesigns have generated proteins that successfully fold to stable, active structures. Some redesigns have been entirely the choice of the investigators, whereas others have incorporated a randomization and selection step to identify optimal sequences. These studies have provided basic guidelines for the rational manipulation of protein structure and stability, they have allowed the dissection of folding pathways and they have generated proteins with the potential for practical therapeutic applications.     

Prevacuolar compartment morphology in vps mutants of Saccharomyces cerevisiae

Over 60 genes have been identified that affect protein sorting to the lysosome-like vacuole in Saccharomyces cerevisiae. Cells with mutations in these vacuolar protein sorting (vps) genes fall into seven general classes based upon their vacuolar morphology. Class A mutants have a morphologically wild type vacuole, while Class B mutants have a fragmented vacuole. There is no discernable vacuolar structure in Class C mutants. Class D mutants have a slightly enlarged vacuole, but Class E mutants have a normal looking vacuole with an enlarged prevacuolar compartment (PVC), which is analogous to the mammalian late endosome. Class F mutants have a wild type appearing vacuole as well as fragmented vacuolar structures. vps mutants have also been found with a tubulo-vesicular vacuole structure. vps mutant morphology is pertinent, as mutants of the same class may work together and/or have a block in the same general step in the vacuolar protein sorting pathway. We probed PVC morphology and location microscopically in live cells of several null vps mutants using a GFP fusion protein of Nhx1p, an Na(+)/H(+) exchanger normally localized to the PVC. We show that cell strains deleted for VPS proteins that have been previously shown to work together, regardless of VPS Class, have the same PVC morphology. Cell strains lacking VPS genes that have not been implicated in the same pathway show different PVC morphologies, even if the mutant strains are in the same VPS Class. These new studies indicate that PVC morphology is another tier of classification that may more accurately identify proteins that function together in vacuolar protein sorting than the original vps mutation classes.     

Proteasomes: protein and gene structures.

Proteasomes are ring- or cylinder-shaped particles that have a sedimentation coefficient of 20S and are composed of a characteristic set of small polypeptides. These particles have a latent multicatalytic proteinase activity. Recently, proteasomes were found to combine reversibly with multiple protein components to form 26S proteolytic complexes that catalyze ATP-dependent, selective breakdown of proteins ligated with ubiquitin. This suggests that the 26S complexes are a new type of ATP-requiring protease in eukaryotic cells. We have studied the structures of various eukaryotic proteasomes at the molecular level by physicochemical and recombinant DNA techniques and have proposed that the gross structures of proteasomes, such as their size and shape, have been highly conserved during evolution. Proteasome subunits appear to be encoded by a family of homologous genes named the "proteasome gene family," which may have evolved from a common ancestral gene. Evidence obtained by genetic analyses in yeast and studies on the levels of proteasome expression in various eukaryotic cells indicates that proteasomes have essential roles in the cell. In this review, we summarize available information on the protein and gene structures of proteasomes and discuss the biological functions of proteasomes.     

Histologic organization of the myocardium of the rat during hypokinesia (stereologic study)

For 30 days Wistar rats have been kept under conditions of hypokinesia. Using stereological methods, the volume and surface density of muscle fibers, microcirculatory bed and connective tissue components have been defined, and volumetric and surface--volume ratios of parenchymal and stromal structures have been calculated. Intracellular stereological parameters of the cardiac myocytes have been studied. Three-dimensional parameters in the parenchymatous-stromal interrelations of the atrophying myocardium accompanied with a deep rearrangement of the intracellular architectonics have been determined, as well as certain destructive changes of the cardiac myocyte ultrastructures. Such a treatment as a decreased functional loading is demonstrated to be the cause of cardiomyocytic lesions.     

Core-coat conductor of lipid bilayer and micromachined silicon

We have etched a groove into a (110) plane of silicon and have covered it with a bilayer of glycerol monooleate. We have varied the depth of the groove, the concentration of salt in the electrolyte and the density of gramicidin in the membrane. We have clamped one end of the groove at a constant voltage with respect to the bath keeping the other end sealed or electrically open with respect to the bath. We have measured (i) the voltage at the center of the groove and at the sealed distal end and (ii) the current through the system in sealed and open configuration. We have found that the spread of voltage is in quantitative agreement with the stationary solutions of Kelvin's equation for a homogeneous cable.     

Chance and the origin of life

Random chemical reactions in the Earth's primitive hydrosphere could have generated no more than 200 bits of information, whereas the first Darwinian organism must have encoded about a million bits, and therefore could not have arisen by chance. This information gap is bridged by separating reproduction from organism, and postulating a reproductive chemical community that would generate information by proto-Darwinian evolution. The information content of the initial community could have been as low as 160 bits, and its evolution might have led to the first Darwinian cell.