Probing Position-Dependent Diffusion in Folding Reactions Using Single-Molecule Force Spectroscopy

Folding of proteins and nucleic acids involves a diffusive search over a multidimensional conformational energy landscape for the minimal-energy structure. When examining the projection of conformational motions onto a one-dimensional reaction coordinate, as done in most experiments, the diffusion coefficient D is generally position dependent. However, it has proven challenging to measure such position-dependence experimentally. We investigated the position-dependence of D in the folding of DNA hairpins as a simple model system in two ways: first, by analyzing the round-trip time to return to a given extension in constant-force extension trajectories measured by force spectroscopy, and second, by analyzing the fall time required to reach a given extension in force jump measurements. These methods yielded conflicting results: the fall time implied a fairly constant D, but the round-trip time implied variations of over an order of magnitude. Comparison of experiments with computational simulations revealed that both methods were strongly affected by experimental artifacts inherent to force spectroscopy measurements, which obscured the intrinsic position-dependence of D. Lastly, we applied Kramers’s theory to the kinetics of hairpins with energy barriers located at different positions along the hairpin stem, as a crude probe of D at different stem positions, and we found that D did not vary much as the barrier was moved along the reaction coordinate. This work underlines the difficulties faced when trying to deduce position-dependent diffusion coefficients from experimental folding trajectories.     

Convenient procedure for the isolation of highly enriched,cytochrome P-450-containing microsomal fraction from Candida tropicalis

The suitability of Ca²⁺ ions for the precipitation of the microsomal fraction from the hydrocarbon-grown yeast Candida tropicalis was evaluated. In the final procedure the microsomes were precipitated by the addition of 16 mm CaCl₂. Crude extracts obtained from cells via spheroplast lysis were centrifuged at 12,000g for 15 min and at 25,000g for 15 min prior to precipitation. The cytochrome P-450 content of the fraction was between 0.22 and 0.35 nmol mg^?1 protein. The isolated microsomes exhibited both hexadecane hydroxylation activity and NADPH-cytochrome c reductase activity.     

Stereoisomeric flavour compounds LXVIII. 2-, 3-, and 4-Alkyl-branched acids,part 2: Chirospecific analysis and sensory evaluation

Enantioselective GC analysis of 4-ethyloctanoic and 4-methylheptanoic acid, using heptakis(2,3-di-O-methyl-6-O-tert-butyldimethylsilyl)-β-cyclodextrin as the chiral stationary phase, is described and the sensory properties of several 4-alkyl-branched acids, using gas chromatography-olfactometry (GC-O) equipment and octakis(2,3-di-O-methyl-6-tert-butyldimethylsilyl)-γ-cyclodextrin as the stationary phase, are evaluated. The chirospecific analysis of various 2-, 3-, and 4-alkyl-branched acids from commercially available Roman chamomile (Chamaemelum nobile (L.) Allioni), Parmesan cheese, and subcutaneous mutton adipose tissue, using either GC-GC (MDGC) or GC-MS analytical methods, is described. © 1994 Wiley-Liss, Inc.     

Über die tagesperiodische Ausdrift von Niphargus aquilex schellenbergi Karaman aus Quellen

Zusammenfassung Die Ausdrift von Niphargus aquilex schellenbergi Karaman aus Quellen verläuft tagesperiodisch. Sie wind durch eine Aufwanderung kompensiert. Drift und Aufwanderung werden als regelmäßige Bewegungen zwischen Spaltenwasser und Quellbach angesehen. Durch Farbmarkierung wird nachgewiesen, daß sie Niphargen unterhalb des Quellabflusses Nahrung aufnehmen und zurückwandern.Extrem große Driftfänge, deren Zusammensetzung sich auch qualitativ von der normalen Drift unterscheidet, können bisher noch night zufriedenstellend erklart werden.Die Tagesperiodik der Ausdrift wird von LD-Wechsel gesteuert. Schwankungen von Temperatur, O₂- und CO₂-Gehalt des Wassers lassen keinen Einfluß erkennen. Es wird vermutet, daß die Tagesperiodik der Ausdrift auf einer Aktivitätsperiodik beruht.

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Summary Amphipodes of the genus Niphargus regularly drift to the surface of springs. The number of animals drifting per 2 hr.-period, shows characteristic changes between day and night. The drift of Niphargus is not an uncontrolled passive process, but likely dependent upon locomotor activity.Dye-markings show, that the drift is compensated by upstream migrations and that Niphargus feeds in the brook before it goes back into the rock crevices.Sometimes the number of Niphargus in the drift-samples is extremly high. In this case the typical pattern of the periodicity and the average size of the amphipodes are markedly changed. This phenomenon (extreme drift) cannot yet be explained satisfactorily.The diel periodicity is timed by LD-changes. Fluctuations of temperature, O₂- and CO₂-content of the water did not produce any effect under natural conditions.

     

Der Nucleolus und seine Beziehungen zu den Ribosomen des Cytoplasmas. Eine Untersuchung an den Malpighischen Gefässen von Drosophila melanogaster

Summary The investigation described herein is concerned with the structure and the behavior of the nucleolus of the cells of the Malpighian tubules of Drosophila melanogaster (see Fig. 16).Primarly the nucleolus consists of ribonucleic-acid granules about 150 Å in diameter which, with electron microscope, are not distinguishable from the ribosomes of the cytoplasm; they lie on or within the structural network of the nucleolus (Fig. 9). During the stages of the life history of the insect the nucleolus shows correspondingly characteristic peculiarities in structure which reflect the varying functional stages of the cell: The structure is loose in the cells of the larva (Fig. 2) and imago (Fig. 12), whereas it is compact in the pupa (Fig. 8, 9). In pupa sufficiently mature for emergence, the nucleolus contains a large central vacuole (Fig. 8, 9). The nucleolus of the larva has a similar appearance after a decrease in assimilation induced by starvation (Fig. 10, 11).There are smaller vacuoles in the nucleolus which mostly are filled with parts of the nucleolus-chromosome. In the pupal stage some of the peripheral constituents of the nucleolus separate and move to the nuclear membrane through which they pass via pores into the cytoplasm (Fig. 3, 4).Larger pieces reach the cytoplasm by local dissolution of the nuclear membrane (Fig. 5). During this process the extruded nucleolar material is dispersed into its structural components. Because the cytoplasm becomes richer in ribosomes during the period of nucleolar extrusion (Fig. 1b), it can be assumed, that the nucleolus must be a source of ribosomes, and that through such an extrusion which has been demonstrated to be a natural process (Fig. 7) the cytoplasm can be provided with ribosomes rapidly.The importance of this phenomenon in the synthesis of imaginal body is discussed.In the nucleus there are many vacuoles with a diameter of about 425 Å (Fig. 15a, b) which pass through the pores of membrane in the cytoplasm (Fig. 15c).

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Die Untersuchung wurde mit Unterstützung der Deutschen Forschungsgemeinschaft durchgeführt.     

Untersuchungen zur entwicklungsgeschichte der araneae [Pardosa Hortensis (Thorell)] unter besonderer berücksichtigung des vorderen prosomaabschnittes

Ohne ZusammenfassungDie Untersuchungen warden mit Unterstützung der Deutschen Forschungsgemeinschaft in den Jahren 1965–1966 im Zoologischen Institut der Universität Kiel durchgeführt. Herrn Prof. Dr. R. Siewing möchte ich meinen herzlichsten Dank sagen für die Überlassung des Themas, für seinen freundlichen Rat und seine stote Hilfsbereitschaft während der Durchführung meiner Untersuchungen. Herrn Dr. G. Scholl bin ich sehr verbunden für zwei mir zur Verfügung gestellte Präparate und eine Zeichnung von Carausius, und ich danke ihm ebenfalls für seine Anteilnahme am Gelingen meiner Arbeit.     

Emerging model systems in evo-devo: horned beetles and the origins of diversity

Horned beetles and beetle horns are emerging as a model system suited to address fundamental questions in evolutionary developmental biology. Here we briefly review the biology of horned beetles and highlight the unusual opportunities they provide for evo-devo research. We then summarize recent advances in the development of new approaches and techniques that are now available to scientists interested in working with these organisms. We end by discussing ways to implement and combine these new approaches to explore new frontiers in evo-devo research previously unavailable to reseachers working outside traditional model organisms.     

Developmental capacitance, genetic accommodation, and adaptive evolution

The concept of genetic accommodation remains controversial, in part because it remains unclear whether evolution by genetic accommodation forces a revolution, or merely a shift in emphasis, in our understanding of how evolution produces adaptive new traits. Here I outline a perspective that largely favors the latter view. I argue that evolution by genetic accommodation can easily be integrated into traditional evolutionary concepts. At the same time, evolution by genetic accommodation invites novel empirical and theoretical approaches that may allow biologists to push the boundaries of our current understanding of the process of evolution and to solve some long-standing controversies. Specifically, I discuss the role of developmental mechanisms as natural, and likely ubiquitous, capacitors of cryptic genetic variation, and the role of environmental perturbations as mechanisms by which such variation can become visible to selection on an individual to population-wide scale. I argue that in combination, developmental capacitance and large-scale environmental perturbations have the potential to facilitate rapid evolution including the origin of novel adaptive features while circumventing otherwise powerful genetic and population-biological constraints on adaptive evolution. I end by highlighting several promising avenues for future empirical research to explore the mechanisms and significance of evolution by genetic accommodation.