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1.
R Glasser  E J Gabbay 《Biopolymers》1968,6(2):243-254
The synthesis of spermine derivatives (II), \documentclass{article}\pagestyle{empty}\begin{document}$ {\rm R}_1 {\rm R}_{\rm 2} {\rm R}_{\rm 3} \mathop {\rm N}\limits^ + \left( {{\rm CH}_2 } \right)_3 \mathop {\rm N}\limits^ + {\rm R}_{\rm 1} {\rm R}_{\rm 2} \left( {{\rm CH}_2 } \right)_2 ]_2 \cdot 4{\rm X}^ - $\end{document}, and spermidine derivatives (III), \documentclass{article}\pagestyle{empty}\begin{document}$ {\rm R}_1 {\rm R}_{\rm 2} {\rm R}_{\rm 3} \mathop {\rm N}\limits^ + \left( {{\rm CH}_2 } \right)_4 \mathop {\rm N}\limits^ + {\rm R}_{\rm 1} {\rm R}_{\rm 2} \left( {{\rm CH}_2 } \right)_3 \mathop {\rm N}\limits^ + {\rm R}_{\rm 1} {\rm R}_{\rm 2} {\rm R}_3 \cdot 3{\rm X}^ - $\end{document}, are reported. The effects of these salts on the helix–coil transition of rA–rU and rI–rC helices were examined. Increasing the size of the hydrophobic substituents, R1, R2, and R3 lowers the degree of stabilization of the helical structure. The disproportionation reaction, 2rA–rU→rA–rU2 + rA occurs readily with salts II and III, especially when the substituents, R1, R2, and R3 are small, i.e., H or Me. Spermine is found to stabilize the rA–rU2 and rI–rC helices to approximately the same extent; however, large differences between the degree of stabilization of rA–rU2 and rI-rC helices are observed when the substituents R1, R2, and R3 are large hydrophobic groups. Similar results are also obtained for the spermidine series. Finally, differences in the interactions of the salts II and III with rA–rU2 and rI–rC helices suggest that the latter helix is denser.  相似文献   

2.
Experimental kinetic data (initial rate and high conversion) on the hydrolysis of cellobiose by 1,4-β-glucosidace (Gliocladium sp.) have been analysed and a competitive inhibition by glucose has been proposed. The determination of kinetic parameters from integral data is based upon algorithms for non-linear optimization and numerical integration. The values of kinetic constants \documentclass{article}\pagestyle{empty}\begin{document}$(v_{\max } = 1.02\frac{{\mu {\rm M}_{{\rm glucose}} }}{{{\rm mg}_{{\rm protein}} \cdot \min }},K_M = 2.6{\rm mM/l, and }K_P = 1.2{\rm mM/l)}$\end{document} agree well with the initialrate results. An important distinction is the confidence limit of parameters. Linear regression analysis shows a virtual accuracy and can lead to wrong conclusions.  相似文献   

3.

Introduction

The Elongator complex, comprising six subunits (Elp1p-Elp6p), is required for formation of 5-carbamoylmethyl (ncm5) and 5-methoxycarbonylmethyl (mcm5) side chains on wobble uridines in 11 out of 42 tRNA species in Saccharomyces cerevisiae. Loss of these side chains reduces the efficiency of tRNA decoding during translation, resulting in pleiotropic phenotypes. Overexpression of hypomodified \( {\text {tRNA}_{{\rm s^{2} {\rm UUU}}}^{{\rm Lys}} , {\rm tRNA}_{{\rm s^{2} {\rm UUG}}}^{{\rm Gln }} \;{\rm and}\;{\rm tRNA}_{{\rm s^{2} {\rm UUC}}}^{{\rm Glu}}} \), which in wild-type strains are modified with mcm5s2U, partially suppress phenotypes of an elp3Δ strain.

Objectives

To identify metabolic alterations in an elp3Δ strain and elucidate whether these metabolic alterations are suppressed by overexpression of hypomodified \( {\text {tRNA}_{{\rm s^{2} {\rm UUU}}}^{{\rm Lys}} , {\rm tRNA}_{{\rm s^{2} {\rm UUG}}}^{{\rm Gln }} \;{\rm and}\;{\rm tRNA}_{{\rm s^{2} {\rm UUC}}}^{{\rm Glu}}} \).

Method

Metabolic profiles were obtained using untargeted GC-TOF-MS of a temperature-sensitive elp3Δ strain carrying either an empty low-copy vector, an empty high-copy vector, a low-copy vector harboring the wild-type ELP3 gene, or a high-copy vector overexpressing \( {\text {tRNA}_{{\rm s^{2} {\rm UUU}}}^{{\rm Lys}} , {\rm tRNA}_{{\rm s^{2} {\rm UUG}}}^{{\rm Gln }} \;{\rm and}\;{\rm tRNA}_{{\rm s^{2} {\rm UUC}}}^{{\rm Glu}}} \). The temperature sensitive elp3Δ strain derivatives were cultivated at permissive (30 °C) or semi-permissive (34 °C) growth conditions.

Results

Culturing an elp3Δ strain at 30 or 34 °C resulted in altered metabolism of 36 and 46 %, respectively, of all metabolites detected when compared to an elp3Δ strain carrying the wild-type ELP3 gene. Overexpression of hypomodified \( {\text {tRNA}_{{\rm s^{2} {\rm UUU}}}^{{\rm Lys}} , {\rm tRNA}_{{\rm s^{2} {\rm UUG}}}^{{\rm Gln }} \;{\rm and}\;{\rm tRNA}_{{\rm s^{2} {\rm UUC}}}^{{\rm Glu}}} \) suppressed a subset of the metabolic alterations observed in the elp3Δ strain.

Conclusion

Our results suggest that the presence of ncm5- and mcm5-side chains on wobble uridines in tRNA are important for metabolic homeostasis.
  相似文献   

4.
Pan BS  Wolyniak CJ  Brenna JT 《Amino acids》2007,33(4):631-638
Summary. Presented here is the first experimental evidence that natural, intramolecular, isotope ratios are sensitive to physiological status, based on observations of intramolecular δ15N of lysine in the mitochondrial mimic Paracoccus denitrificans. Paracoccus denitrificans, a versatile, gram-negative bacterium, was grown either aerobically or anaerobically on isotopically-characterized ammonium as sole cell-nitrogen source. Nitrogen isotope composition of the biomass with respect to source ammonium was = −6.2 ± 1.2‰ for whole cells under aerobic respiration, whereas cells grown anaerobically produced no net fractionation ( = −0.3 ± 0.23‰). Fractionation of 15N between protein nitrogen and total cell nitrogen increased during anaerobic respiration and suggests that residual nitrogen-containing compounds in bacterial cell membranes are isotopically lighter under anaerobic respiration. In aerobic cells, the lysine intramolecular difference between peptide and sidechain nitrogen is negligible, but in anaerobic cells was a remarkable Δ15Np − s = δ15Npeptide − δ15Nsidechain = +11.0‰, driven predominantly by enrichment at the peptide N. Consideration of known lysine pathways suggests this to be likely due to enhanced synthesis of peptidoglycans in the anaerobic state. These data indicate that distinct pathway branching ratios associated with microbial respiration can be detected by natural intramolecular Δδ15N measurements, and are the first in vivo observations of position-specific measurements of nitrogen isotope fractionation.  相似文献   

5.
E J Gabbay 《Biopolymers》1967,5(8):727-747
Information concerning the structures of rA–rU, rA–rU2 rI–rC, rA–rI2, and acid rA helices in solutions is reported. Through the use of diquaternary ammonium salts of the general structure, \documentclass{article}\pagestyle{empty}\begin{document}$ {\rm R}_1 {\rm R}_2 {\rm R}_3 \mathop {\rm N}\limits^ + ({\rm CH}_2 )n\mathop {\rm N}\limits^ + {\rm R}_1 {\rm R}_2 {\rm R}_3 \cdot 2{\rm Br}^ - $\end{document} (I), it is shown that (1) the distances between adjacent negatively charged oxygen atoms on the helix increases in the following order rA–rI2 < rI–rC < rA–rU ? rA–rU2; (2) the density of the helices increases in the order. rA–rI2 < rA–rU < rA–rU2 < rI–rC; (3) there is a large hydrophobia site in rA–rI2 and possibly also in rA–rU, rA–rU2, and rI–rC helices; (4) the results of the interactions between the salts of type I and the helices may be formulated in semi-quantitative terms by the use of two parameters, α, and β which are shown to be related to the charge separation and the density of the helices, respectively; (5) the studies in solutions compare favorably with the x-ray studies on the fibers; and (6) the acid rA helix differs significantly from the other helices by the fact that the electrostatic interstrand interactions between the negatively charged oxygen atom of a phosphate group and the positively charged 10-amino group of adenine contribute significantly to the stabilization of the helix, and thus it is found that the presence of the salts, I, leads to a significant destabilization of the acid rA helix.  相似文献   

6.
Respiration rates are reported to increase exponentially with temperature. Respiration rates of woody tissues are commonly measured as CO2 efflux rates () from that tissue. However, this paper describes clear variations in stem that were not related to temperature for the case of a young beech (Fagus sylvatica L.) and oak (Quercus robur L.) tree during the dormant season. The CO2 concentration ([CO2]) in the xylem of the beech tree showed similar temperature-independent variations. The trees were grown in a growth chamber in which radiation patterns and temperature were kept constant. was measured with an IRGA connected to cuvettes surrounding a stem segment. Xylem [CO2] was measured in situ using a CO2 microelectrode. Depressions in and [CO2] occurred during the light period, despite equal temperatures in the light and dark period. Explanations found in literature for discrepancies in the exponential relationship between temperature and are the influence of (1) sap flow or (2) decreased cell water content. However, (1) the variations were observed in the dormant season, when no sap flow was observed yet, and (2) reduced cell water content was not likely to be apparent as differences in stem transpiration rates between the dark and light period were not significant. Hence, previously formulated theories failed to explain our results. This work therefore provides a new ground for discussion on other possible causes of daytime depressions in . One might be the refixation of respired CO2 by corticular photosynthesis in the stem parts adjacent to the stem segment enclosed by the cuvette.  相似文献   

7.
The transient response method is introduced to elucidate the mechanism of reaction over immobilized enzyme. Glucose oxidation over the glucose oxidase that was immobilized on ion-exchange resin using glutaraldehyde as a linking agent is selected as an example here. The transient responses of a fixed-bed reactor to step increases and decreases in glucose, oxygen, and gluconolactone feed concentrations have been monitored and interpreted. From some responses, we have found that gluconolactone is formed in the reaction of glucose with adsorbed oxygen, while hydrogen peroxide is formed in the reaction of oxygen with adsorbed glucose. Combining all information from interpreting the responses with the literature, a mechanistic picture can be obtained as follows: \documentclass{article}\pagestyle{empty}\begin{document}$$ \begin{array}{*{20}c} {E_{{\rm ox}} + G \to E_{{\rm red}} GL} \\ {E_{{\rm red}} GL \to E_{{\rm red}} + GL} \\ {E_{{\rm red}} + {\rm O}_2 \to E_{{\rm ox}} {\rm H}_2 {\rm O}_2 } \\ {E_{{\rm ox}} {\rm H}_2 {\rm O}_2 \to E_{{\rm ox}} + {\rm H}_2 {\rm O}_2 } \\ \end{array} $$\end{document}.  相似文献   

8.
Summary The energy requirements of Adélie penguin (Pygoscelis adeliae) chicks were analysed with respect to body mass (W, 0.145–3.35 kg, n=36) and various forms of activity (lying, standing, minor activity, locomotion, walking on a treadmill). Direct respirometry was used to measure O2 consumption ( ) and CO2 production. Heart rate (HR, bpm) was recorded from the ECG obtained by both externally attached electrodes and implantable HR-transmitters. The parameters measured were not affected by hand-rearing of the chicks or by implanting transmitters. HR measured in the laboratory and in the field were comparable. Oxygen uptake ranged from in lying chicks to at maximal activity, RQ=0.76. Metabolic rate in small wild chicks (0.14–0.38 kg) was not affected by time of day, nor was their feeding frequency in the colony (Dec 20–21). Regressions of HR on were highly significant (p< 0.0001) in transmitter implanted chicks (n=4), and two relationships are proposed for the pooled data, one for minor activities ( ), and one for walking ( ). Oxygen consumption, mass of the chick (2–3 kg), and duration of walking (T, s) were related as , whereas mass-specific O2 consumption was related to walking speed (S, m·s-1) as .Abbreviations bpm beats per minute - D distance walked (m) - ECG electrocardiogram - HR heart rate (bpm) - ns number of steps - RQ respiratory quotient - S walking speed (m·s-1) - T time walked (s) - W body mass (kg)  相似文献   

9.
Surface free energy (SFE; γ SV) of 16 fruit epicarps present on the Chilean market was calculated by two approaches: the acid–base and Zisman. The results show that the fruit epicarps were low surface energy since the magnitude of γ SV falls within a narrow range, between 37 and 44 mJ m − 2. Zisman approach gave a critical surface tension values, γ cr lower than the SFE calculated by the acid–base approach. Significant differences in SFE between the fruits may be explained by the variation in the chemical composition of epicuticular waxes. The polar (gABSV\gamma^{\rm AB}_{\rm SV}) and apolar (gLWSV\gamma^{\rm LW}_{\rm SV}) components of the SFE were also calculated and a mathematical relation was between both values was found. Values of gABSV\gamma^{\rm AB}_{\rm SV} and gLWSV\gamma^{\rm LW}_{\rm SV} could also be associated with the fruit family and the tissue origins in the ovary region. Finally, it has been shown that fruit epicarps exhibited predominantly electron-donator behaviour since $\gamma_{\rm SV}^- > \gamma_{\rm SV}^+$\gamma_{\rm SV}^- > \gamma_{\rm SV}^+. We believe that the results reported here can potentially impact in food engineering because the compatibility of coatings and fruit epicarps depends on the interaction of their respective chemical and physical properties.  相似文献   

10.
Kinetics of ethanol inhibition in alcohol fermentation   总被引:3,自引:0,他引:3  
The inhibitory effect of ethanol on yeast growth and fermentation has been studied for the strain Saccharomyces cerevisiae ATCC No. 4126 under anaerobic batch conditions. The results obtained reveal that there is no striking difference between the response of growth and ethanol fermentation. Two kinetic models are also proposed to describe the kinetic pattern of ethanol inhibition on the specific rates of growth and ethanol fermentation: \documentclass{article}\pagestyle{empty}\begin{document}$$\begin{array}{*{20}c} {\frac{{\mu _i }}{{\mu _0 }} = 1{\rm } - {\rm }\left( {\frac{P}{{P_m }}} \right);\alpha } \hfill & {\left( {{\rm for}\ {\rm growth}} \right)} \hfill \\ {\frac{{\nu _i }}{{\nu _0 }} = 1{\rm } - {\rm }\left( {\frac{P}{{P'_m }}} \right);\beta } \hfill & {\left( {{\rm for}\ {\rm ethanol}\ {\rm production}} \right)} \hfill \\ \end{array}$$\end{document} The maximum allowable ethanol concentration above which cells do not grow was predicted to be 112 g/L. The ethanol-producing capability of the cells was completely inhibited at 115 g/L ethanol. The proposed models appear to accurately represent the experimental data obtained in this study and the literature data.  相似文献   

11.
The data processing method of the turbidimetric bioassay of nisin was modified to facilitate its industrial application. The influence of the initial indicator concentration was minimized by a redefined specific dose of the bacteriocin as the quotient between the titer of the added bacteriocin and the initial population density of the indicator in the suspension. It was found that d c = 0.125 μg ml−1 was the critical dose of nisin that can cause a complete inhibition of the indicator, Pediococcus acidilactici UL5, with an initial OD of 0.135. To eliminate the interference of the cell debris, an equation, , exploiting d c, was formulated to obtain the intrinsic survival proportion. The use of the specific dose of the bacteriocin and the intrinsic survival proportion as parameters of the dose/response curve greatly enhanced its repeatability and feasibility. A dual-dosage approach was developed to further simplify the conventional standard dose/response curve method.  相似文献   

12.
A method proposed in recent literature was applied to evaluate the average shear rate ( [(g)\dot]av ) \left( {\dot{\gamma }_{\rm av} } \right) in three pneumatic bioreactors of 5-dm3 working volume: bubble column, split airlift, and concentric-tube airlift. The volumetric oxygen transfer coefficient (k L a) is the appropriate characteristic parameter to assess the average shear rate ( [(g)\dot]av ) \left( {\dot{\gamma }_{\rm av} } \right) in this methodology. Correlations for [(g)\dot]av \dot{\gamma }_{\rm av} as a function of superficial gas velocity in the riser region (U GR) and rheological fluid properties (consistency index, K, and flow index, n) were obtained for each model of pneumatic bioreactor studied. The [(g)\dot]av \dot{\gamma }_{\rm av} values estimated by the proposed methodology lay within the range of values calculated by classical correlations. The proposed correlations were utilized to predict the [(g)\dot]av \dot{\gamma }_{\rm av} during the Streptomyces clavuligerus cultivations carried out at the same specific air flow rate (3.5 vvm) in the different types of pneumatic bioreactors. The lowest values of [(g)\dot]av \dot{\gamma }_{\rm av} related to the highest values of consistency index (K) were found for the bubble column bioreactor, and the highest values of [(g)\dot]av \dot{\gamma }_{\rm av} related to the lowest values of K were found for the concentric-tube airlift bioreactor. Intermediate values were found for the split airlift bioreactor. The results showed that high [(g)\dot]av \dot{\gamma }_{\rm av} values affect the structural health of the mycelia by the rupture of the hipha.  相似文献   

13.
A novel NMR pulse sequence is introduced to determine the glycosidic torsion angle χ in 13C,15N-labeled oligonucleotides. The quantitative Γ-HCNCH measures the dipolar cross-correlated relaxation rates (pyrimidines) and (purines). Cross-correlated relaxation rates of a 13C,15N-labeled RNA 14mer containing a cUUCGg tetraloop were determined and yielded χ-angles that agreed remarkably well with data derived from the X-ray structure of the tetraloop. In addition, the method was applied to the larger stemloop D (SLD) subdomain of the Coxsackievirus B3 cloverleaf 30mer RNA and the effect of anisotropic rotational motion was examined for this molecule. It could be shown that the χ-angle determination especially for nucleotides in the anti conformation was very accurate and the method was ideally suited to distinguish between the syn and the anti-conformation of all four types of nucleotides. Electronic Supplementary Material The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

14.
A consecutive, first-order, irreversible, biochemical reaction, \documentclass{article}\pagestyle{empty}\begin{document}$ A{\textstyle{{k(\theta)} \over {{\rm Enzyme }1}}} \to B{\textstyle{{k(\theta)} \over {{\rm Enzyme 2}}}} \to C $\end{document}, taking place in a series of N reactors with product recycle is considered. A discrete version of the maximum principle is used to derive general equations necessary for maximizing the production of (1) the final product, C, by choosing the temperature or the pH value in each reactor, and (2) the intermediate product, B, by choosing the reactor volume. A numerical computation for a series of three reactors with recycle is illustrated. The effects of varying the recycle rates on the optimal state and decision variables are also presented.  相似文献   

15.
Estimation of coefficient of coancestry using molecular markers in maize   总被引:6,自引:0,他引:6  
Summary The coefficient of coancestry (fAB) between individuals A and B is the classical measure of genetic relationship. fAB is determined from pedigree records and is the probability that random alleles at the same locus in A and B are copies of the same ancestral allele or identical by descent (ibd). Recently, the proportion of molecular marker variants shared between A and B (SAB) has been used to measure genetic relationship. But SAB is an upwardly-biased estimator of fAB, especially between distantly-related lines. fAB, SAB, and adjusted (to remove bias) estimates of molecular marker similarity (f AB M ) were compared. RFLP banding patterns at 46 probe-restriction enzyme combinations were obtained for 23 maize inbred lines derived from the Iowa Stiff Stalk Synthetic (BSSS) maize (Zea mays L.) population, and for 4 non-BSSS lines. f AB M was estimated as , where A (or B) was the average proportion of RFLP variants shared between inbred A (or inbred B) and the non-BSSS lines. The average fAB among 253 pairwise combinations of BSSS lines was 0.212, whereas the average SAB was 0.397. The average f AB M was 0.162, indicating that the upward bias in SAB was effectively removed. SAB and fAB were significantly different ( = 0.05) in 76.3% of the comparisons, whereas 24.9% of the f AB M values differed significantly from fAB. The latter result suggests that selection and/or drift were present during inbred line development and that fAB may not be an accurate measure of the true proportion of ibd alleles between two lines. Cluster analyses based on S AB M and f AB M grouped lines according to pedigree, although several exceptions were noted. The presence of shared molecular marker variants between unrelated lines must be considered when setting SAB-based minimum distances for varietal protection. Under simplified conditions, more than 250 molecular marker loci are necessary to obtain sufficiently precise estimates of coefficient of coancestry using molecular markers.A contribution from Limagrain Genetics, a Group Limagrain company  相似文献   

16.
There have been few studies quantifying litterfall, standing litterstock and gross litter decomposition following forest conversion to plantation crops such as cocoa. Additionally, an assessment of changing processes occurring in forest floor litter systems with plantation age is lacking. We investigated litterfall production, standing litter changes and litter decomposition along a chronosequence of shaded cocoa farm fields (secondary forest, 3, 15 and 30-year-old) in the moist semi-deciduous forest belt in the Ashanti Region of Ghana in West Africa over 24 months. Mean annual litterfall production differed significantly among study sites and ranged from 5.0 to 10.4 Mg DM ha?1. Similarly, standing litter differed significantly between land-use /plot ages. The results showed significant differences in quality between litter from forest and litter from cocoa plantations. Litterfall from forests had higher concentrations of nitrogen and lower concentration of soluble polyphenols and lignin compared to litter from cocoa systems. Monthly decomposition coefficients (k) estimated as $ k = {{\left( {{\text{A}} - \left( {{\text{L}}_1 - {\text{L}}_0 } \right)} \right)} \mathord{\left/ {\vphantom {{\left( {{\text{A}} - \left( {{\text{L}}_1 - {\text{L}}_0 } \right)} \right)} {\left( {{{\left( {{\text{L}}_1 + {\text{L}}_0 } \right)} \mathord{\left/ {\vphantom {{\left( {{\text{L}}_1 + {\text{L}}_0 } \right)} 2}} \right. } 2}} \right)}}} \right. } {\left( {{{\left( {{\text{L}}_1 + {\text{L}}_0 } \right)} \mathord{\left/ {\vphantom {{\left( {{\text{L}}_1 + {\text{L}}_0 } \right)} 2}} \right. } 2}} \right)}} $ , where A is litterfall production during the month, L0 is the standing litterstock at the beginning of the month and L1 is the standing litterstock at the end of the month. Annual decomposition coefficients (k L ) were similar in cocoa systems (0.221–0.227) but higher under secondary forests (0.354). Correlations between litter quality parameters and the decomposition coefficient showed nitrogen and lignin concentrations as well as ratios that include nitrogen are the best predictors of decomposition for the litters studied. Our results confirm the hypothesis that decomposition decreases following forest conversion to shaded cocoa systems because of litter quality changes and that decomposition rates correlate to litter quality differences between forest and cocoa ecosystems. The study also showed that standing litter pools and litterfall production in recently converted cocoa plantations are low compared to secondary forests or mature cocoa systems. Management strategies involving the introduction of upper canopy species during plantation development with corresponding replacement of tree mortality with diverse fast growing species will provide high quality and quantity litter resources.  相似文献   

17.
L Yuan  S S Stivala 《Biopolymers》1972,11(10):2079-2089
The effect of dielectric constant (D) of the solvent on the viscosity of heparin was examined using the relation \documentclass{article}\pagestyle{empty}\begin{document}$ \eta _{{\rm sp}} /c = [\eta ]_\infty (1 + k/\sqrt c) $\end{document}, where [η] is the shielded intrinsic viscosity obtained by extrapolating \documentclass{article}\pagestyle{empty}\begin{document}$ \eta _{{\rm sp}} /c\,{\rm vs}{\rm . }\,1/\sqrt c ) $\end{document} to infinite concentration, and k is an interaction parameter independent of the dielectric constant of the solvent. This equation was previously reported by the authors9 for describing the reduced viscosities of strong polyelectrolytes in salt-free polar solvents. It was found that the [η] of heparin increases linearly with increasing dielectric constant of the solvent whereas the k values were, within experimental error, independent of D in the range 54.7 < D < 93.2 examined. Graded hydrolysis of heparin from its acid form (heparinic acid) at 57°C resulted in samples of varying degree of desulfation with corresponding decrease in biological activity. It was found that both [η] and k decrease with increasing desulfation.  相似文献   

18.
A new 3D, spin-state-selective coherence transfer NMR experiment is described that yields accurate measurements for eight scalar or dipolar couplings within a spin system composed of a methylene adjacent to a methine group. Implementations of the experiment have been optimized for proteins and for nucleic acids. The experiments are demonstrated for C–C moieties of the third IgG-binding domain from Streptococcal Protein G (GB3) and for C –C groups in a 24-nt RNA oligomer. Chemical shifts of C, C and H (respectively C , C and H ) are dispersed in the three orthogonal dimensions, and the absence of heteronuclear decoupling leads to distinct and well-resolved E.COSY multiplet patterns. In an isotropic sample, the E.COSY displacements correspond to 1JCH, 2JCH2+2JCH3, 2JCH, 1JCH2+1JCH3, 1JCH22JH2H3, 1JCH32JH2H3, 3JHH2 and 3JHH3 for proteins, and 1J , 2J J , 2J , 1J +1J , 1J J , 1J J , 3J and 3J in nucleic acids. The experiment, based on relaxation-optimized spectroscopy, yields best results when applied to residues where the methine–methylene group corresponds to a reasonably isolated spin system, as applies for C, F, Y, W, D, N and H residues in proteins, or the C –C groups in nucleic acids. Splittings can be measured under either isotropic or weakly aligned conditions, yielding valuable structural information both through the 3J couplings and the one-, two- and three-bond dipolar interactions. Dipolar couplings for 10 out of 13 sidechains in GB3 are found to be in excellent agreement with its X-ray structure, whereas one residue adopts a different backbone geometry, and two residues are subject to extensive 1 rotamer averaging. The abundance of dipolar couplings can also yield stereospecific assignments of the non-equivalent methylene protons. For the RNA oligomer, dipolar data yielded stereospecific assignments for six out of the eight C H2 groups in the loop region of the oligomer, in all cases confirmed by 1J ^{1} $$" align="middle" border="0"> J , and H resonating downfield of H .Supplementary material to this paper is available in electronic form at http://dx.doi.org/10.1007/s10858-005-0175-z.  相似文献   

19.
Release rates of recently fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ from non-exchangeable interlayer sites in 2:1 silicate minerals were determined for decomposed granite (DG) saprolites from three locations in California, USA. Recently-fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ release from the DG substrate was quantified by extracting diffused $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ with H-resin, as well as a native, annual grass Vulpia microstachys. The $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ release data varied with via the method of extraction, which included H-resin pre-treatments (Na+ or H+) and V. microstachys uptake (mycorrhizal inoculated or uninoculated). After 6 weeks (1008 h), more $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ was recovered from fixed interlayer positions by the H-resins as compared to uptake by V. microstachys. The H+ treated H-resins recovered more released $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ (≈94 mg ${\text{NH}}^{{\text{ + }}}_{{\text{4}}} - {\text{N}}\;{\text{kg}}^{1} $ or (12%) of total fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ ) in two of the three DG samples as compared to the Na+ treated resins, (which recovered ≈70–78 mg ${\text{NH}}^{{\text{ + }}}_{{\text{4}}} - {\text{N}}\;{\text{kg}}^{{{\text{ - 1}}}} $ (or 9–10%) of the total fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ ). The V. microstachys assimilated 8–9% of the total fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ with mycorrhizal inoculum as compared to only 2% without a mycorrhizal inoculum, over the same time period. The fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ release kinetics from the H-resin experiments were most accurately described by first order and power function models, and can be characterized as biphasic using a heterogeneous diffusion model. Uptake of both the 15N and ambient, unlabelled N from the soils was closely related to plant biomass. There was no significant difference in percent of N per unit of biomass between the control and mycorrhizal treatments. The findings presented here indicate that observed, long-term $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ release rates from DG in studies utilizing resins, may overestimate the levels of fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ made available to plants and microorganisms. Additionally, the study suggested that mycorrhizae facilitate the acquisition and plant uptake of fixed $ {\text{NH}}^{{\text{ + }}}_{{\text{4}}} $ , resulting in markedly increased plant biomass production.  相似文献   

20.
Hairy root cultures of Gentiana macrophylla were established by infecting the different explants four Agrobacterium rhizogenes strains namely A4GUS, R1000, LBA 9402 and ATCC11325, and hairy root lines were established with A. rhizogenes strain R1000 in 1/2 MS + B5 medium. Initially, 42 independent hairy root clones were maintained and seven clones belongs to different category were evaluated for growth, morphology, integration and expression of Ri T-DNA genes, and alkaloid contents in dry root samples. On the basis of total root elongation, lateral root density and biomass accumulation on solid media, hairy root clones were separated into three categories. PCR and Southern hybridization analysis revealed both left and right T-DNA integration in the root clones and RT-PCR analysis confirmed the expression of hairy root inducible gene. GUS assay was also performed to confirm the integration of left T-DNA. The accumulation of considerable amounts of the root-specific secoiridoid glucosides gentiopicroside was observed in GM1 ( and ) and the GM2 ( and DNA) type clones in considerably higher amount whether as two but callus-type clones (GM3) accumulated much less or only very negligible amounts of gentiopicroside. Out of four media composition the 1/2 MS + B5 vitamin media was found most suitable. We found that initial establishment of root cultures largely depends on root:media ratio. Maximum growth rate was recorded in 1:50 root:media ratio. The maximum biomass in terms of fresh weight (33-fold) was achieved in 1/2 MS + B5 media composition after 35 days in comparison to sixfold increase in control. The biomass increase was most abundant maximum from 15 to 30 days. Influence of A. rhizogenes strains and Ri plasmid of hairy root induction, the possible role of the TL-DNA and TR-DNA genes on growth pattern of hairy root, initial root inoculum:media ratio and effect of media composition is discussed.  相似文献   

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