Peptide mechanics: A force field for peptides and proteins working with entire residues as smallest units

The central ingredient of any structure modeling tool is a molecular model or force field that accounts for proper geometry and energy calculation. For protein and peptide modeling based on entire amino acids as building blocks, we describe a peptide force field in which each amino acid is represented by a single point in space, taken at the position of the α-carbon atom. Apart from the positional coordinates, these units carry the two torsional angles φ and Ψ as additional degrees of freedom to account for the orientations of the peptide links. While some of the energy terms are analogous to expressions in atomic force fields, the presence of the angular variables leads to fundamental differences with new features and additional terms with no atomic counterparts. The force field reproduces secondary structure elements with very good accuracy. Globular parts of tertiary packing stay near the experimental structures with a rms deviation in C-α positions of 0.1–0.3 nm and about 25° in φ and Ψ depending on the size of the structure. A tendency for larger discrepancies is observed in exposed loops or terminal segments the conformations of which may be strongly influenced by neighboring domains. Finally, a scope of possible applications is presented. They range from modeling activities, such as model building by homology, to coarse scanning of conformation space in conformation analysis and structure determination. An extension to a dynamics model would offer the possibility to eliminate the less interesting high-frequency modes that in all-atom force-field dynamics absorb most of the computational effort.     

Catchments as conservation units for riverine biodiversity

The geological structure and longitudinal nature of river systems provide a possible barrier to the dispersal of lotic organisms. This has the potential to drive evolutionary processes such as genetic differentiation and subsequent allopatric speciation. In the conservation of lotic ecosystems population and evolutionary processes have largely been ignored. The traditional approach to river conservation has been focussed toward maintaining the physical habitat that provides the template for the biota. With a shift toward recognising the catchment as the primary operational unit for the conservation and management of lotic systems an examination of the biological relevance of these catchment units is required. This paper examines the structural influence of catchment units over population structure in lotic organisms and the extent to which catchments reflect populations and represent evolutionarily significant units. These being unique assemblages representing segments of biological diversity that share a common evolutionary lineage and contain the potential for a unique evolutionary future. This is of particular importance given the increased use of inter-basin water transfers to move water between historically isolated catchments and recognition of the catchment as the primary unit for the conservation and management of river ecosystems.     

Nuclei act as independent and integrated units of replication in a Xenopus cell-free DNA replication system.

We have used a novel approach to investigate the control of initiation of replication of sperm nuclei in a Xenopus cell-free extract. Nascent DNA was labelled with biotin by supplementing the extract with biotin-11-dUTP, and isolated nuclei were then probed with fluorescein-conjugated streptavidin. Flow cytometry was used to measure the biotin content of individual nuclei and their total DNA content. This showed that incorporation of the biotinylated precursor increases linearly with DNA content. Haploid sperm nuclei replicate fully to reach the diploid DNA content over 2-6 h in the extract. Synthesis stops once the diploid DNA content is reached. Different nuclei enter S phase at different times over greater than 1.5 h, although they share the same cytoplasmic environment. Nuclei reach their maximum rates of synthesis soon after entry into S phase and some replicate fully in less than 0.5 h, resembling the rates of replication observed in the intact egg. These results indicate that initiations are coordinated within each nucleus such that the nucleus is the fundamental unit of replication in the cell-free system.     

New roles for pyruvate kinase M2: working out the Warburg effect

The origins and role of the Warburg effect have remained uncertain for many years. Two recent studies demonstrate that an embryonic- and cancer-cell-specific isoform of the enzyme pyruvate kinase M2 (PKM2) is regulated by binding to phospho-tyrosine motifs and promotes increased cell growth and tumor development. PKM2 enhances the use of glycolytic intermediates for macromolecular biosynthesis and tumor growth. These findings illustrate the distinct advantages of this metabolic phenotype in cancer cell growth.     

Using tea stalk lignocellulose as an adsorbent for separating decaffeinated tea catechins

Lignocelluloses prepared from woody tea stalk, pine sawdust and sugarcane bagasse were used as adsorbents to isolate decaffeinated catechins from tea extracts and compared with synthetic macroporous resin HPD 600. HPD 600 had the highest adsorption capacity to catechins, followed by tea stalk lignocellulose while lignocelluloses of pine sawdust and bagasse the least. Tea stalk lignocellulose absorbed preferentially tea catechins and showed a good selectivity. HPD 600 absorbed caffeine and tea catechins simultaneously. The kinetics data of tea stalk lignocellulose showed a good fit with the Langmuir isotherm model. It is considered that tea stalk lignocellulose is an alternative low-cost adsorbent for preparing decaffeinated tea catechins.     

GPs in principle but not in practice: a study of vocationally trained doctors not currently working as principals.

OBJECTIVES--To identify doctors who are vocationally trained but not currently practising as principals in general practice; their reasons for not practising as principals; and whether the prospect of a re-entry course would appear to this group. DESIGN--Postal questionnaire survey based on semistructured interviews. SUBJECTS--Doctors who had been vocationally trained but were not currently practising as principals: 351 possible subjects identified by a process of "networking." SETTING--Trent Regional Health Authority. RESULTS--166 of the doctors who replied fitted the criteria (100 women; 66 men). The out of hours commitment was ranked as the most important factor for not practising as a principal--95 women and 50 men rated it important--followed by difficulty in combining work with family commitments--84 women, 31 men. 82 respondents (49%) said they would be interested in a re-entry course if one were available. CONCLUSIONS--There is a pool of vocationally trained doctors in Trent region who are not practising as principals in general practice. More flexible working patterns and the availability of a re-entry course could make the post of principal in general practice a more attractive proposition to these doctors.     

Formation of evolutionary approach as an explanatory principle for ecology

Although the connection of ecology with evolutionary idea and specifically with Darwinism was proclaimed for a long time it seems that Herbert Spencer's approach with its emphasize on natural equilibrium was much more often used as its real theoretical base. Elements of Darwinian approach appeared only in 1920-30s in works of those few researchers who studying the distribution and population dynamics of different species tried to understand general mechanisms providing their continuing existence. Later, in the middle of 1950s the first attempts were undertaken to consider the population life history (primarily the age specific schedule of death and reproduction) as a result of natural selection aimed to maintain the necessary level of fitness. A special attention in these studies that burgeoned in 1980-90s was paid to looking for various trade-offs between particular parameters of life history, e.g., between the survival of juveniles and fecundity of adults. The problem of life history optimization became central for the whole branch of science named "evolutionary ecology". Though traditionally this branch is connected with Darwinism, it is rooted rather in Spencer's ideas on moving equilibrium and deals more with static than dynamic. Disproportionately less attention was paid to the evolution of communities since these formations could be hardly interpreted as units of Darwinian selection. Moreover, the ecologists dealing with biosphere as a unified biogeochemical system began insist on "nondarwinian" nature of its evolution. The author considers this opinion as not sufficiently grounded. Darwin's ideas about unavoidable exponential growth, intrinsic for any population, consequent deficiency of resources, and differential survival and reproduction of individuals are still useful while studying the evolution of living organisms (phylogenetics) or the development of biosphere as a global ecosystem.     

Protein structure similarity as guiding principle for combinatorial library design

Proteins are modularly built from a limited set of approximately 1000 structural domains. The evolutionary relationship within a domain family suggests that the knowledge about a common fold structure can be exploited for the design of small molecule libraries in the development of inhibitors and ligands. This principle has been used for the synthesis of inhibitors for kinases sharing the same fold. It can also be applied for proteins which share the same fold architecture yet belong to different functional classes. Bestatin--originally known as an aminopeptidase inhibitor--was employed as guiding structure for the development of leukotriene A4 hydrolase inhibitors. A combinatorial approach helped to identify inhibitors for sulfotransferases which share structural similarity with nucleotide kinases using a kinase inhibitor core structure as guiding principle.     

Evidence for chromosomal replicons as units of sister chromatid exchanges

Chromosomal replicons have been described as the cytological counterpart of DNA replicon clusters and have previously been studied in vitro using premature chromosome condensation-sister chromatid differentiation (PCC-SCD) techniques. Chromosomal replicons are visualized as small SCD segments in S-phase cells, and measurement of these segments can provide estimates of relative chromosomal replicon size corresponding to DNA replicon clusters functioning coordinately in S-phase. Current hypotheses of sister chromatid exchange (SCE) formation postulate that sites of SCE induction are associated with active replicons or replicon clusters. We have applied the PCC-SCD technique to in vivo studies of mouse bone marrow cells that have been treated with cyclophosphamide (CP) for two cell cycles. We have been able to visualize chromosomal replicons, as well as SCEs which have been induced in vivo by CP treatment, simultaneously in the same cells. Chromosomal replicons visualized as small SCD segments were measured in PCC cells classified at early or late S-phase based on SCD segment size prevalence. Early S-phase (E/S) PCC cells contained 90% of the SCD segments measured clustered in a segment size range of 0.1 to 0.8 m with a peak value around 0.3 to 0.6 m regardless of CP treatment. As the cells progressed through S-phase, late S-phase (L/S) PCC cells were characterized by the appearance of larger SCD segments and even whole SCD chromosomes in addition to small SCD segments. A concentration of units around 0.4 to 1.0 m was found for L/S SCD segment size distributions regardless of CP treatment with an apparent bimodal profile. Our in vivo data support the existence of a subunit organization of chromosomal replication with a basic functional unit being 0.3 to 0.6 m in size. In addition, we have found that this chromosomal unit of replication or chromosomal replicon does not seem to be functionally perturbed by the mutagen CP. We also found that small SCD segments of 0.4 to 0.7 m in length were involved in the formation of an SCE, suggesting that both spontaneous and CP-induced SCEs occur between chromosomal replicons. These findings provide direct cytogenetic evidence to support a replicon cluster/chromosomal replicon model for SCE formation.     

Syntheses and studies of amamistatin B analogs reveals that anticancer activity is relatively independent of stereochemistry, ester or amide linkage and select replacement of one of the metal chelating groups

A series of analogs of the amamistatin natural products was designed and synthesized to facilitate additional anticancer structure-activity relationships. The results indicate that the anticancer activity is relatively independent of stereochemistry, ester or amide linkage and replacement of the oxazoline/oxazole based iron-binding group with a catechol.     

The human platelet as an independent unit for sulfate conjugation

The human platelets possess a full complement of enzymes capable of synthesizing N-acetyldopamine (NADA) 35sulfate from ATP, Mg++ and sodium 35sulfate. The pH optimum for this three-step overall sulfate conjugation (comprising of the ATP sulfurylase, APS kinase and phenolsulfotransferase reactions) is 8.6 and the reactions proceeded progressively for several hours. Both ATP and Mg++ ions, above their respective optimal concentrations of 5 and 7 mM, inhibited the sulfate conjugation of NADA. The apparent Km values for NADA as determined by the phenolsulfotransferase (PST) and overall reactions were similar in magnitude being 2.6 and 4.8 microM, respectively, while that for sodium 35sulfate was 202 microM. A comparison of these two activities in 62 platelet preparations of normal subjects showed that the rate of the PST reaction was generally higher than the overall reaction even though the PST assay was carried out at suboptimal concentration of PAPS. There was a positive correlation (r = 0.82) between the two sets of data, suggesting that the PST reaction probably has some control over the rate of overall sulfate conjugation.     

Rubisco as a taxonomic tool: Introducing a simple electrophoretic technique for separating the holoenzyme

A simple electrophoretic technique for separating Rubiscoholoenzyme (ribulose bisphosphate carboxylase-oxygenase) from crude extracts is introduced. No preliminary purification steps are required. To prove the validity of the technique we rely on well studiedBrassica spp., tobacco and spinach. We recommend this method for studies concerned with intrageneric relationships and for proving the intraspecific constancy of Rubisco.