Ice friction during speed skating.

During speed skating, the external power output delivered by the athlete is predominantly used to overcome the air and ice frictional forces. Special skates were developed and used to measure the ice frictional forces during actual speed skating. The mean coefficients of friction for the straights and curves were, respectively, 0.0046 and 0.0059. The minimum value of the coefficient of ice friction was measured at an ice surface temperature of about -7 degrees C. It was found that the coefficient of friction increases with increasing speed. In the literature, it is suggested that the relatively low friction in skating results from a thin film of liquid water on the ice surface. Theories about the presence of water between the rubbing surfaces are focused on the formation of water by pressure-melting, melting due to frictional heating and on the 'liquid-like' properties of the ice surface. From our measurements and calculations, it is concluded that the liquid-like surface properties of ice seem to be a reasonable explanation for the low friction during speed skating.     

The effect of repeated temperature shock on baker's yeast

Summary In a recycle system in which evaporation is used for ethanol recovery during fermentation, temperature changes of the broth in the loop will occur. These repeated temperature shocks may have an effect on the microbial ethanol production rate. In this study such repeated temperature changes were simulated in a recycle system with ethanol production by baker's yeast. The magnitude of the temperature change, as well as the time of exposure to this change were found to have an effect on the ethanol production rate. A temperature increase from 30°C in the fermentor to 35°C or more in the recycle loop led to a significantly lower ethanol concentration in the broth. This effect became negligible at a short exposure time of 18 s of the yeast to the higher temperature. Correspondence to: R. G. J. M. van der Lans     

Mechanisms of intermolecular homologous recombination in plants as studied with single- and double-stranded DNA molecules.

To elucidate the mechanism for intermolecular homologous recombination in plants we cotransformed Nicotiana tabacum cv Petit Havana SR1 protoplasts with constructs carrying different defective derivatives of the NPTII gene. The resulting kanamycin resistant clones were screened for possible recombination products by PCR, which proved to be a valuable technique for this analysis. Our results show that the double-stranded circular DNA molecules used in this study recombine predominantly via a pathway consistent with the single-strand annealing (SSA) model as proposed for extrachromosomal recombination in mammalian cells. In the remaining cases recombination occurred via a single reciprocal recombination, gene conversion and possibly double reciprocal recombination. Since single-stranded DNA is considered to be an important intermediate in homologous recombination we also established the recombination ability of single-stranded DNA in intermolecular recombination. We found that single-stranded DNA enters in recombination processes more efficiently than the corresponding double-stranded DNA. This was also reflected in the recombination mechanisms that generated the functional NPTII gene. Recombination between a single-stranded DNA and the complementing DNA duplex occurred at similar rates via a single reciprocal recombination and the SSA pathway.     

Electrophysiological studies of forskolin-induced changes in ion transport in the human colon carcinoma cell line HT-29 cl.19A: Lack of evidence for a cAMP-activated basolateral K+ conductance

Summary Forskolin (i.e, cAMP)-modulation of ion transport pathways in filter-grown monolayers of the Cl^–-secreting subclone (19A) of the human colon carcinoma cell line HT29 was studied by combined Ussing chamber and microimpalement experiments.Changes in electrophysiological parameters provoked by serosal addition of 10^–5 m forskolin included: (i) a sustained increase in the transepithelial potential difference (3.9±0.4 mV). (ii) a transient decrease in transepithelial resistance with 26±3 · cm² from a mean value of 138±13 · cm² before forskolin addition, (iii) a depolarization of the cell membrane potential by 24±1 mV from a resting value of –50±1 mV and (iv) a decrease in the fractional resistance of the apical membrane from 0.80±0.02 to 0.22±0.01. Both, the changes in cell potential and the fractional resistance, persisted for at least 10 min and were dependent on the presence of Cl^– in the medium. Subsequent addition of bumetanide (10^–4 m), an inhibitor of Na/K/2Cl cotransport, reduced the transepithelial potential, induced a repolarization of the cell potential and provoked a small increase of the transepithelial resistance and fractional apical resistance. Serosal Ba²⁺ (1mm), a known inhibitor of basolateral K⁺ conductance, strongly reduced the electrical effects of forskolin. No evidence was found for a forskolin (cAMP)-induced modulation of basolateral K⁺ conductance.The results suggest that forskolin-induced Cl^– secretion in the HT-29 cl.19A colonic cell line results mainly from a cAMP-provoked increase in the Cl^– conductance of the apical membrane but does not affect K⁺ or Cl^– conductance pathways at the basolateral pole of the cell. The sustained potential changes indicate that the capacity of the basolateral transport mechanism for Cl^– and the basal Ba²⁺-sensitive K⁺ conductance are sufficiently large to maintain the Cl^– efflux across the apical membrane. Furthermore, evidence is presented for an anomalous inhibitory action of the putative Cl^– channel blockers NPPB and DPC on basolateral conductance rather than apical Cl^– conductance.     

Solid-state 13C and 15N NMR study of the low pH forms of bacteriorhodopsin

The visible absorption of bacteriorhodopsin (bR) is highly sensitive to pH, the maximum shifting from 568 nm (pH 7) to approximately 600 nm (pH 2) and back to 565 nm (pH 0) as the pH is decreased further with HCl. Blue membrane (lambda max greater than 600 nm) is also formed by deionization of neutral purple membrane suspensions. Low-temperature, magic angle spinning 13C and 15N NMR was used to investigate the transitions to the blue and acid purple states. The 15N NMR studies involved [epsilon-15N]lysine bR, allowing a detailed investigation of effects at the Schiff base nitrogen. The 15N resonance shifts approximately 16 ppm upfield in the neutral purple to blue transition and returns to its original value in the blue to acid purple transition. Thus, the 15N shift correlates directly with the color changes, suggesting an important contribution of the Schiff base counterion to the "opsin shift". The results indicate weaker hydrogen bonding in the blue form than in the two purple forms and permit a determination of the contribution of the weak hydrogen bonding to the opsin shift at a neutral pH of approximately 2000 cm-1. An explanation of the mechanism of the purple to blue to purple transition is given in terms of the complex counterion model. The 13C NMR experiments were performed on samples specifically 13C labeled at the C-5, C-12, C-13, C-14, or C-15 positions in the retinylidene chromophore. The effects of the purple to blue to purple transitions on the isotropic chemical shifts for the various 13C resonances are relatively small. It appears that bR600 consists of at least four different species. The data confirm the presence of 13-cis- and all-trans-retinal in the blue form, as in neutral purple dark-adapted bR. All spectra of the blue and acid purple bR show substantial inhomogeneous broadening which indicates additional irregular distortions of the protein lattice. The amount of distortion correlates with the variation of the pH, and not with the color change.     

Interpretation of polymorphic DNA patterns in the human alpha-amylase multigene family.

Previous molecular studies have clearly shown that the human amylase locus has a very complicated structure. Multiple salivary and pancreatic amylase genes are present on haplotypes with variable numbers of genes. To study the population heterogeneity, human genomic DNA from family members and random individuals was digested with a number of different restriction enzymes and hybridized with probes representing various parts of the human pancreatic amylase cDNA. The complex patterns obtained were, in most cases, compatible with predictions from the restriction enzyme maps of cloned human amylase genes. With some enzymes deviations from the predicted intensities of the bands associated with the pancreatic amylase gene AMY2A were observed. These findings can be explained by unequal homologous crossovers between AMY2A and AMY1A, resulting in haplotypes with one gene less or one gene more than the haplotypes described thus far. Moreover, a very complicated TaqI polymorphism was found that can be explained by homologous crossovers between different salivary amylase genes. Because some salivary amylase genes have an inverted orientation with respect to the others, these data provide evidence for the occurrence of intrachromosomal, homologous crossovers, as proposed by us previously (P. C. Groot et al., 1990, Genomics 8: 97-105).     

The recombinant congenic strains for analysis of multigenic traits: genetic composition.

The genetic control of susceptibility to many common diseases, including cancer, is multigenic both in humans and in animals. This genetic complexity has presented a major obstacle in mapping the relevant genes. As a consequence, most geneticists and molecular biologists presently focus on "single gene" diseases. To make the multigenic diseases accessible to genetic and molecular analysis, we developed a novel genetic tool, the recombinant congenic strains (RCS) in the mouse (4). The RC strains are produced by inbreeding of mice of the second backcross generation between two inbred strains, one of which serves as the "donor" and the other as the "background" strain. A series of RCS consists of approximately 20 strains, each carrying a different set of genes: approximately 12.5% genes from the common donor inbred strain, the remaining 87.5% from the common background inbred strain. As the set of donor strain genes in each RC strain is different, the nonlinked genes of the donor strain involved in the control of a multigenic trait, e.g., cancer susceptibility, become distributed into different RC strains where they can be analyzed one by one. Hence, the RCS system transforms a multigenic trait into a series of single gene traits, where each gene contributing to the multigenic control can be mapped and studied separately. Recently we demonstrated that the RCS system is indeed capable of resolving multigenic traits, which are hardly analyzable otherwise, by mapping four new colon tumor susceptibility loci (8; P. C. Groot, C. J. A. Moen, W. Dietrich, L. F. M. van Zutphen, E. S. Lander, and P. Demant, unpublished results). For successful application of the RCS system, extensive genetic characterization of the individual recombinant congenic strains is essential. In this paper we present detailed information about the genetic composition of three series of RC strains on the basis of typing of 120-180 markers distributed along all autosomes. The data indicate that the relative representation of the donor strain genes in the RC strains does not deviate from the theoretical expectation, and that the RC strains achieved a very high degree of genetic homogeneity and for all practical purposes can be considered inbred strains. The density and distribution of markers reported here permits an effective mapping of unknown genes of donor strain origin at almost all autosomal locations. Much of this information has been obtained using the new class of genetic markers, the simple sequence repeat polymorphisms.(ABSTRACT TRUNCATED AT 400 WORDS)     

Sequence-tagged sites (STSs) spanning 4p16.3 and the Huntington disease candidate region.

The generation of sequence-tagged sites (STSs) has been proposed as a unifying approach to correlating the disparate results generated by genetic and various physical techniques being used to map the human genome. We have developed an STS map to complement the existing physical and genetic maps of 4p16.3, the region containing the Huntington disease gene. A total of 18 STSs span over 4 Mb of 4p16.3, with an average spacing of about 250 kb. Eleven of the STSs are located within the primary candidate HD region of 2.5 Mb between D4S126 and D4S168. The availability of STSs makes the corresponding loci accessibility to the general community without the need for distribution of cloned DNA. These STSs should also provide the means to isolate yeast artificial chromosome clones spanning the HD candidate region.     

Calcium absorption by fish intestine: The involvement of ATP-and sodium-dependent calcium extrusion mechanisms

Summary Measurements of unidirectional calcium fluxes in stripped intestinal epithelium of the tilapia,Oreochromis mossambicus, in the presence of ouabain or in the absence of sodium indicated that calcium absorption via the fish intestine is sodium dependent. Active Ca²⁺ transport mechanisms in the enterocyte plasma membrane were analyzed. The maximum capacity of the ATP-dependent Ca²⁺ pump (V _m :0.63 nmol·min^–1 mg^–1,K _m : 27nm Ca²⁺) is calculated to be 2.17 nmol·min^–1·mg^–1, correcting for 29% inside-out oriented vesicles in the membrane preparation. The maximum capacity of the Na⁺/Ca²⁺ exchanger with high affinity for Ca²⁺ (V _m :7.2 nmol·min^–1·mg^–1,K _m : 181nm Ca²⁺) is calculated to be 13.6 nmol·min^–1·mg^–1, correcting for 53% resealed vesicles and assuming symmetrical behavior of the Na⁺/Ca²⁺ exchanger. The high affinity for Ca²⁺ and the sixfold higher capacity of the exchanger compared to the ATPase suggest strongly that the Na⁺/Ca²⁺ exchanger will contribute substantially to Ca²⁺ extrusion in the fish enterocyte. Further evidence for an important contribution of Na⁺/Ca²⁺ exchange to Ca²⁺ extrusion was obtained from studies in which the simultaneous operation of ATP-and Na⁺-gradient-driven Ca²⁺ pumps in inside-out vesicles was evaluated. The fish enterocyte appears to present a model for a Ca²⁺ transporting cell, in which Na⁺/Ca²⁺ exchange activity with high affinity for Ca²⁺ extrudes Ca²⁺ from the cell.