Foam behavior of biological media

Summary The surface tension and foaminess of (a) unlimited, (b) substrate limited, and (c) oxygen transfer limited growth media of Hansenula polymorpha were measured using methanol, ethanol or glucose as a substrate.The time dependence of can be described by the Avrami-Überreiter relationship: log (2.3 log V)=n log t+log b, where V = (_O – _eq/(_t – _eq, and _O, _t and _eq are at t_M=0, t_M=t and t_M (equilibrium value).The constants n and b are functions of the fermentation time t_F as long as the growth is unlimited but they are constant in the state of limited growth. With glucose substrate, the foaminess can be presented as a definite function of the time, t_DG, which is necessary to attain _eq. With alcohol as a substrate no definite (t_DG) function was found.Symbols b constant in Eq. (1) - n constant in Eq. (1) - S substrate concentration - T temperature - t_M time h (measured from the beginning of the determination of the surface tension ) - t_F cultivation time h (measured from the time of inoculation) - t_DG time (min) necessary to attain the equilibrium surface tension ) - X dry biomass concentration (gl^–1) - V (_O – _eq)/(_t – _eq) - V_S equilibrium volume of the foam (cm³) - V_G volumetric gas flow rate during the estimation of (cm³ s^–1) - vvm volumetric gas flow rate with regard to the volume of the medium (min^–1) - w_SG superficial gas velocity (cm s^–1) - _m maximum specific growth rate (h^–1) - V_S/V_G foaminess (s) - surface tension, mMm^–1 (milli Newton m^–1) - _O at t_M=0 - _eq equilibrium surface tension ( at t_M) - _t at t_M=t - HP probes from Hansenula polymorpha cultivation - NLG non limited growth - OTLG oxygen transfer limited growth - SLG substrate limited growth     

Membrane pathways for water and solutes in the toad bladder: II. Reflection coefficients of the water and solute channels

Summary Urea and water transport across the toad bladder can be separately activated by low concentrations of vasopressin or 8 Br-cAMP. Employing this method of selective activation, we have determined the reflection coefficient () of urea and other small molecules under circumstances in which the bladder was transporting urea or water. An osmotic method for the determination of was used, in which the ability of a given solute to retard water efflux from the bladder was compared to that of raffinose (=1.0) or water (=0). When urea transport was activated (low concentration of vasopressin), for urea and other solutes was low, (_urea,0.08–0.39;_acetamide, 0.55; _{ethylene glycol}, 0.60). When water transport was activated (0.1mm 8 Br-cAMP) _urea approached 1.0 _urea also approached 1.0 at high vasopressin concentrations. In a separate series of studies, _urea was determined in the presence of 2×10^–5 m KMnO₄ in the luminal bathing medium. Under these conditions, when urea transport is selectively blocked, _urea rose from a value of 0.12 to 0.89. Thus, permanganate appears to close the urea transport channel. These findings indicate that the luminal membrane channels for water and solutes differ significantly in their dimensions. The solute channels, limited in number, have relatively large radii. They carry a small fraction (approximately 10%) of total water flow. The water transport channels, on the other hand, have small radii, approximately the size of a water molecule, and exclude solutes as small as urea.     

A method for the estimation of gene flow parameters from a population structure caused by restricted gene flow and genetic drift

Summary A method has been developed which enables the estimation of the plant gene flow parameters _p (pollen dispersal), _s (seed dispersal) and t (outcrossing rate) from a selection-free continuously structured population in equilibrium. The method uses Wright's F-coefficients and introduces a new F-function which describes the genetic similarity as a function of the spatial distance. The method has been elaborated for wind pollinated plant species but can be modified for insect pollination and for animal species. In practice allozymes will provide for the necessary neutral genetic variation. The more loci used and the more intermediate the gene frequencies, the more reliable the results. For the estimation of _p and t together (when the outcrossing rate is not known) at least two chromosomally unlinked loci are required. The method for estimating _s depends on whether the plant species is annual or perennial. The mechanism of selfing has been analysed by the explanation of the value of t by three components: population density (d), pollen flow (_p) and relative fertilization potential of own pollen (Z). The concepts of neighbourhood size and isolation by distance, developed by Wright, who used a single gene flow parameter , have been extended to the situation which is realistic for seed plants, using all three parameters _p, _s and t. When _p is large with respect to _s, _s largely determines the value of the neighbourhood size, whereas _p is the most dominating factor in isolation by distance. The use of local effective population size and mean gene transport per generation instead of neighbourhood size and neighbourhood area, respectively, is proposed to avoid confusion. Computer simulations have been carried out to check the validity and the reliability of the method. Populations of 200 plants, using two or three loci with intermediate allele frequencies, gave good results in the calculation of _p with known value of t and of _s and N_e. With unknown t, especially with lower values of t, larger populations of at least 1,000 plants are necessary to obtain reasonably accurate results for _p and mean gene transport per generation M.Grassland Species Research Group Publication No. 81     

RNA polymerase sigma-related proteins in Escherichia coli: detection by antibodies against a synthetic peptide

Summary Antibodies were raised against a synthetic tetradecameric peptide with an amino acid sequence, DLIQEGNIGLMKAV, which corresponds to the most highly conserved region of bacterial RNA polymerase factors. In a Western-blot analysis of total Escherichia coli proteins, the antiserum reacted specifically with at least three proteins with apparent molecular weights of 75 kDa, 27 kDa and 23 kDa, in addition to the known factors (⁷⁰ and ³²). The majorities of ⁷⁰ and ³² were recovered as associated forms with the RNA polymerase on glycerol gradient centrifugation, while the other cross-reacting proteins were not. Unambiguous evidence was obtained which indicated that the intracellular level of ³² increased rapidly upon heatshock, at least in the strain containing high copy numbers of the rpoH gene.     

Theoretical studies on the necessary number of components in mixtures

Summary Theoretical studies on the optimal numbers of components in mixtures (for example multiclonal varieties or mixtures of lines) have been performed according to phenotypic yield stability (measured by the parameter variance). For each component i, i = 1, 2,..., n, a parameter u_i with 0 u_i 1 has been introduced reflecting the different survival and yielding ability of the components. For the stochastic analysis the mean of each u_i is denoted by u ₁ and its variance by _i ² For the character total yield the phenotypic variance V can be explicitly expressed dependent on 1) the number n of components in the mixture, 2) the mean of the _i ² 3) the variance of the _i ² 4) the ratio and 5) the ratio _i ² /² where denotes the mean of the u _i and _u ² is the variance of the u _j. According to the dependence of the phenotypic stability on these factors some conclusions can be easily derived from this V-formula. Furthermore, two different approaches for a calculation of necessary or optimal numbers of components using the phenotypic variance V are discussed: A. Determination of optimal numbers in the sense that a continued increase of the number of components brings about no further significant effect according to stability. B. A reduction of b % of the number of components but nevertheless an unchanged stability can be realized by an increase of the mean of the u _i by 1% (with and _u ² assumed to be unchanged). Numerical results on n (from A) and 1 (from B) are given. Computing the coefficient of variation v for the character total yield and solving for the number n of components one obtains an explicit expression for n dependent on v and the factors 2.-5. mentioned above. In the special case of equal variances, _i ² = _o ² for each i, the number n depends on v, x = (₀/)² and y = (_u/)². Detailed numerical results for n = n (v, x, y) are given. For x 1 and y 1 one obtains n = 9, 20 and 79 for v = 0.30, 0.20 and 0.10, respectively while for x 1 and arbitrary y-values the results are n = 11, 24 and 95.This publication is an extended version of a lecture given at the 1984-EUCARPIA meeting (Section Biometrics in Plant Breeding) in Stuttgart-Hohenheim (Federal Republic of Germany)     

Amber mutations in the structural gene for RNA polymerase sigma factor of Escherichia coli

Summary Amber mutants of Escherichia coli K-12 affected in the structural gene (rpoD) for th subunit of RNA polymerase have been obtained from a strain harboring a temperature-sensitive amber suppressor (supF-Ts6) which is active only at low temperatures. These mutants grow normally at low temperature (30°C) but do not grow at high temperature (42°C) due to the inability to synthesize factor. In one mutant studied in detail (rpoD40), the rate of -factor synthesis at 30°C is about half that of the wild type and is decreased to 10%–15% within 1 h of incubation at 42°C. The synthesis of core polymerase subunits or bulk protein is virtually unaffected at least for 2 h. The defect of the mutant in synthesis and growth at high temperature can be suppressed by any of the amber suppressors tested (supD, supE or supF). RNA-polymerase holoenzymes prepared from the mutant cells carrying each of the suppressors (grown at 42°C) exhibit different thermostabilities attributable to alterations in the factor. The reduced synthesis in the mutant is accompanied by the synthesis of polypeptide tentatively identified as amber fragment. These results as well as the genetic mapping data indicate that the amber mutation (rpoD40) resides within the structural gene for the factor and directly affects synthesis upon inactivation of the suppressor at high temperature.     

A membrane-bound enzyme complex synthesising glucan and glucomannan in pine tissues

Particulate membrane preparations isolated from cambial cells and differentiating and differentiated xylem cells of pine (Pinus sylvestris L.) trees synthesised [¹⁴C]glucans using either guanosine 5-diphosphate (GDP)-D-[U-¹⁴C]glucose or uridine 5-diphosphate (UDP)-D-[U-¹⁴C]glucose as glycosyl donors. Although these glucans had -(13) and -(14) linkages in an approximate ratio 1:1, the distribution of the linkages in the glucan synthesised from GDP-D-glucose was different from that synthesised from UDP-D-glucose. The synthesis of the mixed -(13) and -(14) glucan from GDP-D-[U-¹⁴C]glucose was changed to that of -(14) glucomannan in the presence of increasing concentrations of GDP-D-mannose. The glucan formed from UDP-D-[U-¹⁴C]glucose was not affected by any concentration of GDP-D-mannose. The membrane preparations epimerized GDP-D-glucose to GDP-D-mannose; however, the low amount of GDP-D-mannose formed was not incorporated into the polymer becaus the affinity of the synthase for GDP-D-glucose was much greater than that for GDP-D-mannose. The glucan formed from GDP-D-glucose and the glucomannan formed from GDP-D-glucose together with GDP-D-mannose were characterized. The apparent K _m and V _max of the glucan synthase for GDP-D-glucose were 6.38 M and 5.08 M·min^-1, respectively. No lipid intermediates were detected during the synthesis of either glucan or glucomannan. The results indicated that an enzyme complex for the formation of the glucomannan was bound to the membrane.Abbreviations GDP guanosine 5-diphosphate - GLC gasliquid chromatography - UDP trridine 5-diphosphate     

Distance-constraint approach to protein folding. I. Statistical analysis of protein conformations in terms of distances between residues

A statistical analysis of protein conformations in terms of the distance between residues, represented by their C atoms, is presented. We consider four factors that contribute to the determination of the distanced _i,i+k between a given pair ofith and(i+k)th residues in the native conformation of a globular protein: (1) the distancek along the chain, (2) the size of the protein, (3) the conformational states of theith to(i+k)th residues, and (4) the amino acid types of the and(i+k)th residues. In order to account for the dependence on the distancek along the chain, the statistics are taken for three ranges, viz., short, medium, and long ranges (k8; 9k20; andk21; respectively). In the statistics of short-range distances, a mean distanceD _k and its standard deviationS _k are calculated for each value ofk, with and without taking into account the conformational states of all residues fromi toi+k (factors 1 and 3). As an Appendix, the relations for converting from the distances between residues into other conformational parameters are discussed. In the statistics of long-range distances, a reduced distanced* _ij (the actual distance divided by the radius of gyration) is used to scale the data so that they become independent of protein size, and then a mean reduced distanceD _l (a, a) and its standard deviation _l (a, a) are calculated for each amino acid pair (a, a) (factors 2 and 4). The effect of the neighboring residues along the chain on the value of the distanced* _ij is explored by a linear regression analysis between the actual reduced distanced* _ij and the mean value over theD _l for all possible pairs of residues in the two segments of the (i–2)th to the (i+2)th and the (j–2)th to the (j+2)th residues. The effect is assessed in terms of the tangentA _l (a, a) of the calculated regression line for each amino acid pair (a, a). In the statistics of medium-range distances, only factors 1 and 4 are considered, to simplify the analysis. The scaled distanced _i,i+k =(d _i,i+k -D _k )/S _k is used to eliminate the dependence onk, the distance along the chain. The propertiesD _m (a, a), _m (a, a) andA _m (a, a) corresponding toD _l (a, a), _l (a, a), andA _l (a, a), and also calculated for each amino acid pair (a, a). The results are interpreted as follows: the smaller values ofD _l (a, a) andD _m (a, a) indicate a preference of the pair (a, a) for a contact (e.g., pairs between hydrophobic amino acids, and pairs of Cys with aromatic amino acids), and the larger values of these quantities indicate a preference for distant mutual location (e.g., pairs between strong hydrophilic amino acids); the smaller values of _l (a, a) and _m (a, a) indicate a strong preference for either contact or noncontact (e.g., pairs between hydrophobic amino acids, and pairs between strong hydrophobic and hydrophilic amino acids, respectively), and the larger values of these quantities indicate the ambivalent/neutral nature of the preference for contact and noncontact (e.g., pairs containing Ser or Thr); the smaller values ofA _l (a, a) andA _m (a, a) indicate that the distance of an (a, a) pair is determined independently of the amino acid character of the neighboring residues along the chain (e.g., some pairs of Cys or Met with other amino acids) and the larger values of these quantities indicare that such amino acid character contributes strongly to the determination of the distance (e.g., pairs containing Ser or Thr, and pairs between amino acids with small side chains). The difference between the statistics for the long- and medium-range distances is also discussed; the former reflect the difference between the hydrophobic and hydrophilic character of the residues, but the latter cannot be easily interpretable only in terms of hydrophobicity and hydrophilicity. The data analyzed here are used in the optimization of an object function to compute protein conformation in a subsequent paper.     

Precision of neural timing: effects of convergence and time-windowing

We study the improvement in timing accuracy in a neural system having n identical input neurons projecting to one target neuron. The n input neurons receive the same stimulus but fire at stochastic times selected from one of four specified probability densities, f, each with standard deviation 1.0 msec. The target cell fires if and when it receives m inputs within a time window of msec. Let _n,m, denote the standard deviation of the time of firing of the target neuron (i.e. the standard deviation of the target neuron's latency relative to the arrival time of the stimulus). Mathematical analysis shows that _n,m, is a very complicated function of n, m, and . Typically, _n,m, is a non-monotone function of m and and the improvement of timing accuracy is highly dependent of the shape of the probability density for the time of firing of the input neurons. For appropriate choices of m, , and f, the standard deviation _n,m, may be as low as . Thus, depending on these variables, remarkable improvements in timing accuracy of such a stochastic system may occur.     

Probabilities of negative estimates of genetic variances

Summary The probability of negative analysis of variance estimates of genetic variance components due to sampling error (Ps) was investigated. The objectives were to evaluate the magnitude of Ps, to compare Ps for estimates of _A ² and _D ² , and to compare Ps for genetic variance component estimates from the nested and factorial mating designs. Ps was defined in terms of ratios of mean squares and the F distribution was used to calculate probabilities of the negative estimates. The results indicated that Ps is often greater than 0.20 for _D ² . It is generally lower for _A ² than for _D ² , and lower for the factorial mating design than the nested mating design.Technical Contribution No. 2589 from the South Carolina Agricultural Experiment Station, Clemson University     

Properties of components of covariance of inbred relatives and their estimates in a maize population

Summary Properties of three parameterizations, denoted as the C-model, D-model and Q-model, for covariances of inbred relatives under assumptions of no linkage or epistasis are explored and compared. Additive variance in an inbred population with inbreeding coefficient F, ² _AF =(1+F) ² _A where ² _A is additive variance in a panmictic population, if Q-model parameters Q _xx and Q _xy are both zero. Conditions sufficient for this to hold are presented in terms of gene frequencies and dominance contrasts (homozygotes vs. heterozygotes). Some other properties and potential uses of estimates of components in the models are also discussed. Estimates of components in the D-model and Q-model were calculated from a maize (Zea mays L.) study from which estimates of components in the C-model were previously published. Of particular interest were the covariance (Q _xy ) of effects of alleles at complete homozygosity with inbreeding depression effects, the covariance (D ₁) of additive effects at panmixia with inbreeding depression effects and the within-locus variance (D ₂, alias Q _xx ) of inbreeding depression effects. Estimates of Q _xy , D ₁, and D ₂ were small and nonsignificant in most cases. For ear height in the second year of the study, D ₂ appeared to be a major component. In some cases, results were obtained which had contradictory implications (negative D ₂ coupled with positive Q _xy or D ₁, and positive D ₂ coupled with negative ² _D ). A negative estimate of one or the other of ² _D or ² _A was obtained in one of the two within-year analyses for every character. Problems in getting realistic results were thought to be owing to excessive multicollinearity among the coefficients of the components in the expectations of the covariances of the kinds of relatives included in the study. Implications for future studies of this kind are discussed.Journal Article No. 87-3-14 of the Kentucky Agricultural Experiment Station published with the approval of the Director     

Study ofO-sialylation of glycoproteins in C6 glioma cells treated with retinoic acid

When treated with retinoic acidin vivo, C6 glioma cells show an enhancement of CMP-Neu5Ac:Gal 1–3 GalNAc-R -2,3 sialyltransferase activity. A 300kDa glycoprotein was detected by lectin affinoblotting in retinoic acid-treated C6 cells which stained weakly or not at all in control cells. Comparative studies with different lectins demonstrated that this glycoprotein contains 2,3 Neu5Ac Gal-GalNAc O-glycan moieties. Cultures in the presence of an inhibitor of O-glycan synthesis (N-acetylgalactosaminide -O-benzyl) demonstrated that enhancement of staining of the 300 kDa glycoprotein was not due to the increase of the 2,3 sialyltransferase but to thede novo synthesis of the polypeptide chain of this glycoprotein.Abbreviations RA retinoic acid - Neu5Ac N-acetylneuraminic acid - CMP-Neu5Ac cytidine 5 monophosphosialate - 2,3 ST CMP-Neu5Ac:Gal 1–3 GalNAc-R -2,3 sialyltransferase - GalNAc-O-benzyl N-acetylgalactosaminide -O-benzyl - Gal1-3GalNAc-O-benzyl Galactosyl 1-3N-acetylgalactosaminide -O-benzyl - TBS Tris-HCl buffer 50mm pH 7.5 containing NaCl 0.15m and Tween 20 0.05% - B1 buffer TBS containing MgCl₂ 1mm, MnCl₂ 1mm and CaCl₂ 1mm     

Stimulation of σ1 Receptors Alleviates Reperfusion-Induced Myocardial Stunning

In vivo or in vitro administration of a ₁ receptor agonist d-SKF 10.047 (1 mg/kg intravenously or 10 mg/l in vitro) promoted an increase in the resistance of isolated perfused rat heart to ischemia/reperfusion injury. Both in vivo and in vitro stimulation of receptors prevents the development of reperfusion contracture and creatine kinase release and increases the developed pressure, double product, +dP/dt, and –dP/dt in the left ventricle. Activation of receptors has no significant effect on the occurrence of reperfusion arrhythmias ex vivo. Stimulation of cardiac sigma receptors is proposed to prevent myocardial stunning.     

A comparative study of variation in codon 33 of the<Emphasis Type="Italic"> rpoS</Emphasis> gene in<Emphasis Type="Italic"> Escherichia coli</Emphasis> K12 stocks: implications for the synthesis of σ<Superscript>s</Superscript>

The Escherichia coli rpoS gene encodes an RNA polymerase sigma factor (sigma S or ^S) required for the expression of stationary-phase genes. In the first published rpoS sequence from E. coli K-12 codon 33 is given as CAG. However, several subsequent independent studies found the amber codon TAG at this position ( rpoSAm). Besides this amber codon, other codons such as TAT have also been found at this location in rpoS. Comparative genome analysis now leads us to propose TAG as the parental codon 33 in rpoS in E. coli K-12. Five different stocks of the strain W3110, which differ in the levels of ^S protein they express, were investigated. We sequenced the rpoS gene from these, and found a T at nucleotide position 97 in four out of the five stocks and a G at position 99 in three out of the five. W1485, a parental strain of W3110, and W3350, a derivative of W3110, are also rpoSAm mutants. Such rpoSAm mutants would be expected to show no RpoS activity. The retention of partial or intermediate ^S activity by suppressor-free rpoSAm mutants is therefore puzzling. We propose that a functional, N-terminally truncated, ^S (1–53^S) can be translated from a Secondary Translation Initiation Region (STIR) located downstream of the amber codon 33. It has recently been reported that a fragment of RpoS (1–53^S) that lacks the first 53 amino acids is functional when synthesized in vivo. Taken together, our results support the hypothesis that the original codon 33 of the rpoS gene in E. coli K-12 strains is the amber codon TAG.Communicated by W. Goebel     

Ionic relations of aeroponically-grown olive genotypes,during salt stress

Two olive (Olea europaea L.) genotypes, Frantoio and Leccino, were exposed to increasing concentrations of NaCl (0-30-60-120 mM) in an aeroponic cultivation system for 60 days. Dry weights and sodium and potassium contents of apical and basal leaves, new and old wood, and roots were measured to determine Na uptake rate, Na translocation rate and K-Na selectivity ratio (S_K,Na). Frantoio showed a higher salt resistance than Leccino. Frantoio and Leccino had a similar Na uptake rate, but largely differed for Na translocation to the shoot. Furthermore Frantoio exhibited a higher K-Na selectivity than Leccino at both whole plant level and above all at the level of shoot system. Resistance mechanism of Frantoio is probably related to Na esclusion by roots and to the ability to maintain an appropriate K/Na ratio in actively growing tissues.Research supported by National Research Council of Italy, Special project RAISA.     

Observations on phase-locking within the response of primary muscle spindle afferents to pseudo-random stretch

In order to uncover encoder properties of primary muscle spindle afferent fibers, time coupling (phase-locking) of action potentials on cyclic muscle stretch was studied by means of pseudo-random noise. In cats Ia action potentials were recorded from dorsal root filaments and the gastrocnemius muscles of one hind leg were stretched. The stimulus time course was a determined sequence of randomly varying muscle length which could be applied repeatedly (sequence duration 0.6 or 20 s). The noise amplitude (standard deviation of displacements) was varied between 5 and 300 m, the upper cut-off frequency of noise f _c was varied between 20 and 100 Hz. The responses to the consecutive pseudo-random noise cycles were displayed as raster diagrams and cycle histograms. Phaselocking characterized the responses at all noise amplitudes outside the near threshold range (>10 m). The higher and f _c , the stronger was the phase-locking of impulses on the stretch. When and f _c were selected to achieve high mean stretch velocities of about 500 mm/s, phase-locking was as precise as 0.15 ms, measured as the variability of spike occurrences with respect to stretch. The rasters obtained with low noise amplitudes (<40 m) showed a loose phase-locking and this gave insight into underlying mechanisms: The elicitation of action potentials caused by dynamic stretch can be prevented by a post-spike depression of excitability. This disfacilitation was very effective in counteracting weak stretch components within the random sequence and less effective or even missing when relatively strong stretch components could force the spike elicitation. This led to the reestablishment of phase-locked patterns. The results were discussed in relation to the known encoder models.     

Genetic architecture of a heterotic cross between two populations of maize

Summary The nature and magnitude of variability in the interpopulation cross of Mezcla Amarillo Selection (MAS), an introduction from CIMMYT, Mexico, and J607, a population developed in India using indigenous, American, and Yugoslavian germplasm, were studied. Interpopulation progenies developed by following the North Carolina Design I were evaluated at two locations. The additive genetic variance component in interpopulation cross, _A(12) ² , and in one population assuming the other population as tester, _A12 ² and _A21 ² were significant for all the traits evaluated, namely ear length, ear girth, kernel rows and days to silk, with one exception. For kernel rows, the dominance variance component, _A(12) ² , was also significant but it was smaller than _A(12) ² . The variance component due to dominance X location interaction, _DL(12) ² , was significant for all traits except kernel rows. In the case of ear length and ear girth, _DL(12) ² was greater than the other components. _AL(12) ² , _AL12 ² and _AL21 ² were not significant for any trait. Expected genetic advance indicated a superiority of half-sib reciprocal recurrent selection over full-sib reciprocal recurrent selection.     

Studies of the action of ceramide-like substances (d- andl-PDMP) on sphingolipid glycosyltransferases and purified lactosylceramide synthase

We have studied the effects ofD-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (D-PDMP) and itsL-enantiomer on glycosphingolipids in cultured normal human kidney proximal tubular cells. We found thatD-PDMP exerted a concentration-dependent reduction in the metabolic labelling and cellular levels of glucosylceramide (GlcCer), lactosylceramide (LacCer), and the globo-series glycosphingolipids, GbOse₃Cer and GbOse₄Cer. It also directly inhibited the activity of UDP-glucose:ceramide 1 4-glucosyltransferase (GlcT-1) and UDP-galactose: GlcCer 1 4 galactosyltransferase (GalT-2). In contrast,L-PDMP had opposite effects on the metabolic labelling of GlcCer, LacCer, and GbOse₃Cer. The levels of GlcCer and LacCer were increased, while the labelling and level of GbOse₄Cer were strongly reduced. Purified GalT-2 from human kidney was inhibited byD-PDMP and stimulated byL-PDMP. It appears likely that the different glycosphingolipid glycosyltransferases possess similar binding sites for the ceramide moiety, which are blocked by binding toD-PDMP and, in the case of GbOse₄Cer synthase, byL-PDMP as well. The stimulatory effects ofL-PDMP on GlcCer and LacCer synthases may be the result of binding to a modulatory site on the glycosyltransferases; in intact cells, the enzyme-analog complex may afford protection against the normal catabolic inactivation of the enzymes.Abbreviations GalT-2 UDP-galactose:GlcCer -galactosyltransferase - GbOse₃Cer Gal1 4Gal1 GlcCer - GbOse₄Cer GalNAc1 3Gal1 4Gal1 GlcCer - GlcCer glucosylceramide - GlcT-1 UDP-glucose:ceramide -glucosyltransferase - GSLs glycosphingolipids - LacCer lactosylceramide - PDMP threo-1-phenyl-2-decanolyamino-3-morpholino-1-propanol     

Free sigma subunit of Bacillus subtilis RNA polymerase binds to DNA

Summary The affinity of Bacillus subtilis RNA polymerase and subunits to DNA was examined by a non-denaturing polyacrylamide slab gel electrophoresis method which made it possible to resolve DNA-bound and free subunits. The results revealed that subunit, but not subunit had a relatively high affinity for double stranded DNA. The subunit was bound maximally to super-coiled pGR1-3 plasmid DNA at a mass ratio of /DNA of 0.7. With B. subtilis double stranded linear DNA one subunit was bound per approximately 1,000 base pairs. The -DNA complex was sufficiently stable for isolation by a molecular gel filtration column. The subunit had much higher affinity for super-coiled than for linear pGR1-3 DNA or for linear double stranded or denatured DNA from B. subtilis, E. coli, and calf thymus. These results indicate that the free B. subtilis subunit, in contrast to the E. coli subunit, can bind by itself to DNA.