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51.
Marco L. F. Giuseppin Hendrikus M. J. van Eijk Marja Hellendoorn José W. van Almkerk 《Applied microbiology and biotechnology》1987,27(1):31-36
Summary The changes in cell wall strength of Hansenula polymorpha have been investigated in continuous cultures with respect to the recovery of methanol oxidase (MOX). Cultures grown on several substrate mixtures that enable induction of MOX have been compared with cultures grown on methanol as the sole inducer. The effects of dilution rate (D) on lysis properties have been studied. The cell wall strength was consistently influenced by growth media and D. Media containing glycerol/methanol showed the slowest lysis kinetics, with a large fraction of non-degradable cell wall material. In continuous cultures grown on a mixture of glucose and methanol both the resistance to zymolyase and the mean cell wall thickness increased at D<0.1 h–1. The yield of MOX by zymolyase lysis is reproducible and up to 100% higher than that of the standard ultrasonic treatment. The lysis kinetics indicated that zymolyase punctures the cell wall; since the release rate of MOX is lower than that of protein, the cell contents will leak through. At D-values>0.2 h–1, both protein and MOX release rates increase, reflecting a change in lysis mechanism due to the increased fraction of thin daughter cells. Kinetic analysis of zymolyase lysis using both physical and enzymatic methods provides information for achieving optimal recovery of MOX.Abbreviations and symbols
C
MOX
MOX activity [MOX units·g X–1]
-
D
dilution rate [h–1]
- MOX
methanol oxidase
- kc
decay rate constant of A 610 nm [min–1]
- kd
decay constant of MOX activity [min–1]
- kdis
dissociation rate constant [min–1]
- kMOX
release rate constant of MOX activity [min–1]
- kp
release rate constant of protein [min–1]
- R
recovery efficiency of enzyme [-]
-
St
stability of enzyme [-] 相似文献
52.
A. De Marco A. M. Petros R. A. Laursen M. Llinás 《European biophysics journal : EBJ》1987,14(6):359-368
The interaction of the isolated human plasminogen kringle 4 with the four -amino acid ligands -aminocaproic acid (ACA), N-acetyl-l-lysine (AcLys), trans-aminomethyl(cyclohexane)carboxylic acid (AMCHA) and p-benzylaminesulfonic acid (BASA) has been further characterized by 1H-NMR spectroscopy at 300 and 600 MHz. Pronounced high-field shifts, reaching 3 ppm, are observed for AMCHA resonances upon binding to kringle 4, which underscores the relevance of ligand lipophilic interactions with aromatic side chains at the binding site. Ligand titration curves for the nine His and Trp singlets found in the kringle 4 aromatic spectrum reveal a striking uniformity in the kringle response to the various ligands. The average binding curves exhibit a clear Langmuir absorption isotherm saturation profile and the data were analyzed under the assumption of one (high affinity) binding site per kringle. Equilibrium association constants (K
a
) and first order dissociation rate constants (k
off) were derived from linearized expressions of the Langmuir isotherm and of the spectral line-shapes, respectively. The results for the four ligands, at 295 K, pH* 7.2, indicate that: (a) AMCHA exhibits the strongest binding (K
a
=159 mM
-1) and ACA the weakest (K
a
=21 mM
–1) with AcLys and BASA falling in between; (b) ACA dissociates readily (k
off = 5.3 × 103 s–1) and AMCHA associates the fastest (k
off = 2.0 × 108
M
–1 s–1) while the kinetics for BASA exchange is relatively slow (k
off = 0.8 × 103 s–1, k
on = 0.6 × 108
M
–1s–1); (c) the ligand-binding kinetics is close to diffussion-controlled.Abbreviations ACA
-aminocaproic acid
- AcLys
N-acetyl-l-lysine
- AMCHA
t-aminomethyl(cyclohexane)carboxylic acid
- BASA
p-benzylaminesulfonic acid
- K4
kringle 4
- NOE
nuclear Overhauser effect
- ppm
parts-per-million
- pH*
glass electrode pH reading uncorrected for deuterium isotope effects
-
K
a
ligand-kringle 4 equilibrium association constant
-
k
off
ligand-kringle 4 dissociation rate constant
-
k
on
ligand-kringle 4 association rate constant 相似文献
53.
B. Friebe M. C. Cermeño F. J. Zeller 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1987,73(3):337-342
Summary C-banding patterns and nucleolar activity were analyzed in Dasypyrum villosum, its added chromosomes to hexaploid wheat and the hexaploid amphiploid Triticum dicoccum-D. villosum. Two different populations of the allogamous species D. villosum (2n= 14, VV) from Greece and Italy were analyzed showing a similar polymorphism for C-banding pattern. Six of the seven addition lines were identified by their characteristic C-banding pattern. No polymorphism between both members of each added alien chromosome was found. Furthermore, nucleolar activity and competition were studied by using silver staining procedure. In D. villosum only one chromosome pair, A, was found to be responsible for organizing nucleoli. The results obtained in the amphiploid and in the addition lines demonstrate that nucleolar activity is restricted to SAT-chromosomes 1B and 6B of wheat, while those of D. villosum remain inactive. 相似文献
54.
M H Boyer B Cami J P Chambost M Magnan J Cattanéo 《European journal of biochemistry》1987,162(2):311-316
55.
The interaction of fibrinogen with the mannose-specific lectins concanavalin A (ConA), its acetyl derivative (Ac-ConA) and Lens culinaris agglutinin (LcH) was studied. Both ConA and LcH interact specifically with individual fibrinogen B beta and gamma chains and with denatured fragments D and E. However, analysis of the binding data shows that four moles of Ac-ConA are bound per mole of fibrinogen with two sets of binding sites (Kd1 = 2.4 microM and Kd2 = 16.6 microM; n1 = n2 = 2) while only two moles of LcH are bound per mole of fibrinogen (Kd = 2.6 microM). Ultracentrifugation studies are also in agreement with the presence in the fibrinogen molecule of two and four binding sites for LcH and Ac-ConA, respectively. No aggregates of fibrinogen formed through LcH or Ac-ConA linkages are observed. The use of a crosslinking reagent and ultracentrifugal analysis of the lectin-fibrinogen fragments D1 and E complexes indicated that ConA, as well as Ac-ConA, interact with both fragments D and E while LcH interacts only with fragment D. Furthermore, the binding of ConA to both D and E domains in the intact fibrinogen molecule is clearly demonstrated by using a bifunctional reagent. The bivalent character of ConA tetramers may be misinterpreted as a lack of accessibility of the lectin to two of the four carbohydrate chains of fibrinogen. The differential binding of LcH and ConA to the carbohydrate chains of fibrinogen can be related to a different exposure of the oligosaccharide in D and E fragments and domains and to the different requirements of both lectins for their binding to glycoproteins. 相似文献
56.
Interaction of epidermal growth factor with micelles monitored by photochemically induced dynamic nuclear polarization-1H NMR spectroscopy 总被引:1,自引:0,他引:1
K H Mayo A De Marco E Menegatti R Kaptein 《The Journal of biological chemistry》1987,262(31):14899-14904
Photochemically induced dynamic nuclear polarization (CIDNP)-1H-NMR spectroscopy has been used to study the interaction of the protein hormone epidermal growth factor (EGF) with micelles of sodium dodecyl sulfate (SDS) and dodecylphosphorylcholine (DPC). Conventional 1H-NMR spectra show that most protein resonances remain unperturbed when micelles are added to solution, which argues that the overall protein conformation is maintained in the presence of SDS or DPC at the concentrations used. Photo-CIDNP enhancements of resonances assigned to aromatic side chains of residues at the COOH terminus and beta-sheet regions of murine EGF (i.e. Trp-49, Trp-50, and Tyr-37) are considerably reduced in the presence of micelles, while resonances of aromatic side chains of residues found elsewhere on the protein surface are mostly unaffected. This suggests that the primary interaction between murine EGF and the micelle occurs at the micelle-bulk solvent interface. The overall negatively charged surface of SDS micelles tends to induce a stronger interaction with the protein compared to the zwitterionic DPC micelles, probably due to electrostatic interactions. Cleavage of the COOH-terminal pentapeptide containing both tryptophan residues enhances the already present, but weak, interaction with Tyr-10 and attenuates it with Tyr-37. A similar interaction pattern is found with rat EGF suggesting that at least concerning these two species of EGF the interaction is somewhat specific and conserved. A simple mass-action model for protein-micelle interaction is also presented. 相似文献
57.
A novel group of very long chain polyenoic fatty acids in dipolyunsaturated phosphatidylcholines from vertebrate retina 总被引:10,自引:0,他引:10
M I Avelda?o 《The Journal of biological chemistry》1987,262(3):1172-1179
Dipolyunsaturated phosphatidylcholines from bovine retina contain a whole series of unusual fatty acids. Methyl esters from these acids are very strongly retained on polar and nonpolar gas-liquid chromatography stationary phases. On thin layers of silica-AgNO3, they separate as tetra-, penta-, and hexaenoic fatty acid methyl esters. After hydrogenation, the three polyunsaturated fractions give the same series of saturated methyl esters, having 20 (or 22)-36 carbon atoms. High pressure liquid chromatography, as well as gas-liquid chromatography, indicates that the new components of the three fractions are even-carbon homologs of well known polyenoic fatty acids of the n-6 and n-3 families, since they behave as series of 20-36-carbon tetraenoic (n-6), pentaenoic (n-3 and n-6), and hexaenoic (n-3) fatty acids. Their occurrence in phospholipid molecules also having docosahexaenoate (22:6) explains the separation of major dipolyunsaturated phosphatidylcholines from retina into dodecaenoic, undecaenoic, and decaenoic fractions after argentation thin layer chromatography. Using high pressure liquid chromatography, the latter are resolved into individual species having 10-12 double bonds and 42-58 carbon atoms. The unusual PCs are thus endowed not only with the highest degree of unsaturation, but with the longest hydrocarbon chains yet reported for vertebrate glycerophospholipids. It is shown that phosphatidylcholines containing the novel fatty acids are highly concentrated in photoreceptor membranes and that they occur in the retina of vertebrates so distant in evolution as fish, birds, and various mammals. 相似文献
58.
Alvaro García-Tejedor A. Raul Castaño Federico Morán Francisco Montero 《Journal of molecular evolution》1987,26(4):294-300
Summary The most relevant properties of hypercycles were previously studied mainly from a theoretical point of view. We have developed a Monte Carlo method simulating hypercyclic organization to obtain information about the dynamics of this prebiotic organization. Nucleation, growth, and selective properties have been tested and the results obtained are in good agreement with those of the theoretical predictions. The influence of hypercyclic organization of the error threshold has also been studied. As a consequence of the emergence of a hypercycle, the value of this threshold decreases. The amount of this decrease depends on the population size. Moreover, for some interval of quality factor values, either the hypercycle organization or an error catastrophe can be produced, depending on the initial conditions. The influence of these phenomena on both the dynamic behavior and evolutionary advantages of the hypercycle, as well as their decisive roles on genome size, are discussed.Presented at the FEBS Symposium on Genome Organization and Evolution, held in Crete, Greece, September 1–5, 1986 相似文献
59.
(1-14C)-labeled (n-6) eicosatetraenoate, (n-3) docosapentaenoate and (n-3) docosahexaenoate (20:4, 22:5 and 22:6, respectively) are efficiently taken up and actively esterified into the lipids of bovine retina after 2 h incubation. Photoreceptor membranes, mitochondria, microsomes and postmicrosomal supernatants, which display significant differences in phospholipid and fatty acid compositions, are isolated after such incubations to study the labeling of lipids. The lipid classes preferentially labeled with the acids (1) largely differ among and within subcellular fractions, while (2) some common features in the treatment of the three polyenes are observed in each fraction. In all of them, the three acids are actively incorporated in phosphatidylcholine; ethanolamine glycerophospholipid, phosphatidylserine (PS) and phosphatidylinositol (PI) are highly labeled with 22:6, 22:5 and 20:4 respectively; within ethanolamine glycerophospholipid, the three label phosphatidylethanolamine in preference to plasmenylethanolamine. Most of the 14C esterified in mitochondria is in phospholipids. The endoplasmic reticulum produces in addition highly labeled triacylglycerols, also found in cytosol. High levels of 14C-labeled diacylglycerols are observed exclusively in photoreceptor membranes, where the specific radioactivity of PI is very high. The total amounts of 14C incorporated (1) are in general similar within a given fraction for the three polyenes, but (2) largely differ among fractions. The labeling of the highly unsaturated phospholipids of photoreceptor membranes is the lowest, while the postmicrosomal supernatant (whose lipids are relatively the poorest in polyenoic fatty acids) contains most of the labeled lipids isolated from retinas under these conditions. The results indicate that polyunsaturated species of retina phospholipids undergo an active synthesis and turnover, as well as an intense intracellular traffic among membranes. 相似文献
60.
The effect on fibrinogen coagulation and fibrinolysis of the mannose-specific lectins concanavalin A, its acetyl derivative and Lens culinaris agglutinin was studied. Concanavalin A and acetyl-concanavalin A, which bind to the four carbohydrate chains of fibrinogen, and L. culinaris agglutinin, which only binds to the carbohydrate present in fibrinogen D domains, has the same effect on the coagulation rate: an inhibition at low lectin concentrations and an increase at high concentrations. On the other hand, L. culinaris agglutinin does not alter fibrin crosslinking while acetyl-concanavalin A produces a slight inhibition of both gamma-gamma and alpha-polymer formation. However, this effect is very small when compared with the clear inhibitory effect produced by concanavalin A. Concanavalin A and acetyl-concanavalin A have an inhibitory effect on the rate of fibrin clot lysis proportional to the lectin concentration. Nearly 100% inhibition was obtained when two lectin-binding sites were occupied by either concanavalin A or acetyl-concanavalin A. However, L. culinaris agglutinin has a clearly weaker effect and more than 50% inhibition was not observed. The comparative study of the effect of the three lectins on fibrinolysis as well as on the formation of fibrinogen aggregates suggests that the inhibitory effect of concanavalin A and acetyl-concanavalin A is primarily due to their binding to the carbohydrate chains of fibrinogen E domain. 相似文献