Effects of growth condition on the structure of glycogen produced in cyanobacterium Synechocystis sp. PCC6803

Growth and glycogen production were characterized for Synechocystis sp. strain PCC6803 grown under continuous fluorescent light in four variations of BG-11 medium: either with (G+) or without (G−) 5 mM glucose, and with a normal (N+, 1.5 g sodium nitrate/L) or a reduced (N−, 0.084 g sodium nitrate/L) nitrogen concentration. Glucose-supplemented BG-11 with a normal nitrogen concentration (N+G+) produced the highest growth rate and the greatest cell density. Although the maximum cell mass production was observed in the N+G+ medium, the highest glycogen yield (19.0 mg/g wet cell mass) was achieved under the glucose-supplemented, nitrogen-limiting condition (N−G+). The addition of glucose enhanced cell growth, while nitrogen limitation apparently directed carbon flux into glycogen accumulation rather than cell growth. Transmission electron microscopic analysis showed that, under nitrogen-limiting conditions (N−G+), glycogen particles accumulated in large amounts and filled the cytosol of the cells. Analysis by high-performance size-exclusion chromatography further revealed that the glycogen produced in N−G+ medium had the longest average branch chain-length (DP10.4) among the conditions tested. When the yield and structure of glycogen were examined in different growth phases, the greatest yield (36.6 mg/g wet cell mass) and the longest branch chain-length (DP10.7) were observed 2 days after the fully grown cells in the N+G+ medium were transferred to the growth restricting (N−G+) medium.     

Reassessment of the random coil conformation: vibrational CD study of proline oligopeptides and related polypeptides.

The "random coil" conformational problem is examined by comparison of vibrational CD (VCD) spectra of various polypeptide model systems with that of proline oligomers [(Pro)n] and poly(L-proline). VCD, ir and uv CD spectra of blocked L-proline oligopeptides [(Pro)n, n = 2-12] in different solvents are reported and compared to the spectra of poly(L-proline) II, poly(L-glutamic acid), and unblocked proline oligomers. Based on the chain-length dependence of the VCD and electronic CD (ECD) spectra of proline oligomers, it is established that VCD spectra are dominated by short-range interactions. The VCD of random coil model polypeptides is shown to be identical in shape but smaller in magnitude than poly(L-proline) II and of similar magnitude to that of (Pro)n (n = 3, 4). Based on the spectral evidence, it is concluded that the "random coil" conformation has a large fraction of helical regions, conformationally similar to the left-handed, 3(1) polyproline II helix, as was previously suggested by Krimm and co-workers. This conclusion is further supported by studies of effects of salt (CaCl2, LiBr, LiClO4), temperature (5-75 degrees C), and pH on the VCD spectra of L-proline oligomers, poly(L-proline) II, and poly(L-glutamic acid). These show that, after each of these perturbations, a significant local ordering remains in the oligomers and polymers studied, and that charged polypeptides such as poly(L-glutamic acid) are more flexible than are polyproline or even L-proline oligomers.     

The glycogen content in stressed marine bivalves: The initial absence of a decrease

• 1.1. Changes in the glycogen content, condition, stomach content and acetic acid concentration of mussels Mytilus edulis and cockles Cerastoderma edule were followed during periods of up to 14 days of exposure (to air) at temperatures of 5 and 20°C.
• 2.2. In animals with a high glycogen content the glycogen is not used during the first 3 to 7 days, at high and low temperature respectively.
• 3.3. After this latent period the glycogen concentration often decreased, coinciding with a high mortality and an increase of the concentration of acetic acid.
• 4.4. In cockles with a low glycogen content, and kept at a high temperature, glycogen can be used from the beginning of the stress period.
• 5.5. Between species no clear differences were found.
• 6.6. The stomach content decreased during exposure; however, the stomach content amounted to only 0.5 to 0.7% of the body weight, and is thought to be of minor importance as an energy source during the stress period.
• 7.7. Especially at the higher temperatures glycogen finally is transformed into acetic acid.
• 8.8. It is concluded that during exposure, the animals do not die because of a lack of energy reserves, but because of a high accumulation of acids.

     

Intrachain end-to-end charge transfer interaction in oligosarcosines

Oligosarcosines with two to seven sarcosine units terminated by an electron donor (p-dimethylaminoanilide group) and an electron acceptor (3,5-dinitrobenzoyl group) were synthesized by a liquid- or solid-phase method. The oligomers showed a distinct charge-transfer (CT) absorption in chloroform and ethanol solutions at low enough concentrations to eliminate the contribution of intermolecular CT interaction. Fractions of the oligomers which form the intrachain CT complex were evaluated at infinite dilution and plotted as a function of the number of sarcosine units n. The plot showed an alternating higher and lower tendency with the increase of n from n = 3 to 7; the total maximum was found at n = 4. The maximum reached 0.24 in chloroform, which is higher than that expected for an unperturbed oligosarcosine chain free from the end-to-end interaction, by an approximate factor of 50. The alternating chain-length dependence was found to be governed by an alternating tendency in the conformational enthalpy required for cyclization.     

Anti-cooperative interactions in single-strand oligomers of deoxyriboadenylic acid

Optical rotatory dispersion measurements were made on the deoxyribo nucleotides d(pA)₂, d(pA)₄, d(pA)₆ and poly(deoxyriboadenylic acid) at neutral pH over the temperature range 5–80°C. and were compared to similar data for the analogous oligoriboadenylic acids. The data were interpreted in terms of a temperature-dependent stacking of the bases in the single-strand deoxyribo oligomers. The thermal transition curves show an inverted chain-length dependence compared to the ribo oligomer curves. These results are explained by a theory of anti-cooperative interaction, where the nucleation parameter σ is >1. The theory, based on a one-dimensional Ising model involving both attractive nearest-neighbor and repulsive next-nearest-neighbor interactions, predicts the inverse chain length dependence and agrees rather well with the experimental data. At and above the transition temperature, the deoxyribo polymer is seen to consist of isolated stacked base pairs separated by at least one unit of random coil, there being only a very small probability for the existence of sequences of stacked residues longer than one. The partition function is seen to undergo an irregular behavior as a function of chain length because of the anti-cooperative phenomenon. It is necessary to use an enthalpy of stacking of ?5.0 kcal./mole in order to fit the experimental data with the theory. This value, 1.5 kcal./mole more positive than the ΔH found for the ribo oligomers, is reasonable, since the 2′ hydroxyl group would be expected to stabilize the stacking interaction in the ribo oligomers. Various kinds of distribution functions are calculated and plotted graphically for this theoretical model. A physical rationale is presented for the use of a repulsive next-nearest-neighbor term in this theory for the deoxyribo oligomers.     

The effects of chain length on the secondary structure of oligoadenylates

The oligoadenylates (Ap)_2–4A have been studied by proton magnetic resonance (pmr) spectroscopy. All the exterior base protons and a number of the interior base proton resonance have been assigned. The results of this work showed that the adenine bases in these oligoadenylates are intramolecularly stacked at 20°C with their bases oriented preferentially in the anti conformation about their respective glycosidic bonds. The oligomers were found to associate extensively even at concentrations of 0.02 M, primarily via “end-to-end” stacking. With increasing temperature, the oligomer bases destack, but it is argued that this unfolding process cannot be described in terms of a two-state stacked versus unstacked model. Instead, the temperature dependences of the base proton chemical shifts support a base-oscillation model. The relationship between this model and the two-state model is discussed. Finally, on the basis of the chain-length dependence of the proton chemical shifts of the various adenine bases, it was concluded that subtle variations in the secondary structure of these oligomers exist with increasing chain length. Evidence is presented to show that the effects of distant base shielding are considerably smaller than what was previously estimated. The observed departures from the “extended dimer” model are attributed to differences in the relative orientations of the bases with respect to their neighbors in the oligomer.     

Comparison of the conformational dynamics of the (1----4)- and (1----6)-linked alpha-D-glucans using 13C-NMR relaxation.

The conformational dynamics of alpha-(1----4)- and alpha-(1----6)-glucan homooligomers in the nanosecond time domain have been compared by measuring the 13C-nmr longitudinal relaxation times T1 for carbons of the terminal and interior sugar residues. Measurements are reported on monomeric glucose and on oligomers containing up to ten glucose residues at room temperature in aqueous solution at concentrations of 3 and 20 g/dL. The carbons of terminal residues display longer relaxation times than do those of interior residues, presumably as a consequence of a greater degree of conformational mobility of the chain ends. The T1s of the reducing terminal residues of all oligomers are significantly longer than those of the corresponding nonreducing termini, a phenomenon that we associate tentatively with the anomeric equilibrium at the reducing end. Carbons of the reducing terminal residues in the beta-anomeric form relax more slowly than their alpha-anomeric counterparts. At 20 g/dL the mean T1s for carbons of the terminal and interior residues attain asymptotic behavior with increasing chain length at a chain length of about six residues, and carbons of the alpha-(1----4)-linked maltooligomers relax significantly more slowly than those of the corresponding alpha-(1----6)-linked isomaltooligomers. The T1s of both glucan series increase with decreasing concentration. This concentration dependence disappears below 3 g/dL, where the T1s of the two series of homoligomers are no longer distinguishable. This suggests that in dilute aqueous solution at room temperature viscous damping effects predominate over contributions to the T1-sensitive conformational dynamics from structural differences in the glycosidic linkage region. At 3 g/dL the approach to long chain-length asymptotic behavior is more protracted than at 20 g/dL, and the T1s of carbons of interior oligomeric residues appear to match the corresponding high-polymer behavior at a chain length of eight and greater.     

Studies of the effect of the sugars ribose, xylose and fructose on the retrogradation of wheat starch gels by x-ray diffraction

The effects of the sugars ribose, xylose and fructose on the retrogradation of wheat starch gels were investigated by measuring the area under the strong 0·516 nm diffraction peak (characteristic of B-type crystalline retrograded starch) as a function of storage time for a series of gels containing different amounts of added sugars. Retrogradation was monitored as the increase in peak area with storage time. The results obtained suggested that all three sugars altered crystallisation and hence retrogradation of the gels. For the concentration regimes studied, xylose and ribose acted by progressively reducing crystallisation with increasing sugar concentration. In the case of fructose two effects were noted. The fructose led to an increase in both thermally reversible and thermally irreversible crystallisation upon storage. For xylose and ribose the increase in crystallisation upon storage was almost totally thermoreversible suggesting that the retrogradation upon storage was dominated by amylopectin crystallisation.     

Control of glycogen synthase and phosphorylase by insulin in rat adipocytes which is unrelated to the concentration of cyclic AMP

Incubation of adipocytes in glucose-free medium with adrenocorticotrophic hormone, epinephrine, isoproterenol, or norepinephrine increased the concentration of cyclic AMP and the percentage of phosphorylase a activity, and decreased the percentage of glycogen synthase I activity. Glucose was essentially without effect on glycogen synthase or phosphorylase in either the presence or absence of epinephrine. Although glucose potentiated the action of insulin to activate glycogen synthase, the hexose did not enhance the effectiveness of insulin in the presence of epinephrine. Likewise, glucose did not increase the ability of insulin to oppose the activation of phosphorylase by epinephrine.The activation of glycogen synthase by insulin was not associated with a decrease in the concentration of cyclic AMP. Insulin partially blocked the rise in cyclic AMP due to isoproterenol, adrenocorticotrophic hormone, and norepinephrine. The maximum effects of isoproterenol on glycogen synthase and phosphorylase were observed when the concentration of cyclic AMP was increased twofold. However, insulin clearly opposed the changes in enzyme activity produced by isoproterenol (and also adrenocorticotrophic hormone, epinephrine and norepinephrine) even though concentrations of cyclic AMP were still increased three- to fourfold. Nicotinic acid opposed the increases in cyclic AMP due to adrenocorticotrophic hormone, isoproterenol and norepinephrine to the same extent as insulin; however, nicotinic acid was ineffective in opposing the activation of phosphorylase and inactivation of glycogen synthase produced by these agents. Thus, it is unlikely that the effects of insulin on glycogen synthase and phosphorylase result from an action of the hormone to decrease the concentration of cyclic AMP.     

Rat liver glycogen metabolism in the perinatal period

The correlation between blood glucose levels, the concentration of glycogen, the activities of glycogen sythase and phosphorylase and their respective kinases and phosphatases was examined in liver of rat fetuses between day 18 of gestation and one day after birth. Between day 18 and 21 there is a rapid increase in the concentration of glycogen and in the activity of synthase a and a much slower increase in the activity of phosphorylase a. The activity of the respective kinases increased rapidly during this period and reached maximun on day 21. The activity of synthase phosphatase and phosphorylase phosphatase increased after day 18, to reach a maximum on day 19 and 20, respectively, but decreased again towards day 21. The possibility that the changes in glycogen concentration and enzyme activities were related to an effect of glucose of AMP on the respective phosphatases was considered. It was found that the Km of phosphatase for glucose in the prenatal period was 5–7 mM, as in the adult. Since the level of blood glucose during this period was constant (2.8 mM), an effect of glucose on phosphatase activity seems unlikely. AMP concentration increased between day 18 and 21 from 6–15 nmol/g. In view of the low level of phosphorylase a activity during this period, the increase in AMP concentration is not considered to be important in the regulation of glycogen breakdown at this time.Immediately after birth blood glucose levels dropped to 5 mg/dl. This was accompanied by a rapid decrease in glycogen concentration and in the activity of glycogen synthase and a rise in phosphorylase activity. Blood glucose levels returned to the initial level within 1 h after birth, whereas the changes in glycogen concentration and enzyme activities continued for at least 3 h after birth. On day 22 all parameters examined had reached the level found in adult rat liver.It is suggested that the rapid changes observed immediately after birth are due to an effect of hypoglycemia mediated by hormones and cannot be ascribed to direct effects of metabolites on the enzyme systems involved.     

Kinetics of binding of oligosaccharides to a homogeneous pneumococcal antibody: dependence on antigen chain length suggests a labile intermediate complex.

Temperature-jump experiments were performed with di-, tetra-, and hexasaccharides derived from type III pneumococcal polysaccharide using a homogeneous corresponding antibody IgG 45-394. A decrease in stability of the oligosaccharide-antibody complexes with decreasing chain length was observed and entirely reflected in the decrease of the association rate constants which were 1.7 X 10(4) M-1 s-1 for the di-, 3.7 X 10(5) M-1 s-1 for the tetra-, and 1.1 X 10(6) M-1 s-1 for the hexasaccharide at 23 degrees C. The dissociation rate constants for all oligomers were about 12 s-1. This marked chain-length dependence of the association rate constants as well as their low values are unexpected for a single binding step. A mechanism is proposed which consists of a fast formation of a labile oligosaccharide-antibody precomplex followed by a slow isomerization step which is induced by the oligosaccharide ligands but which is chain-length independent.     

Production and Characterisation of Cross-Linked Enzyme Crystals (Clecs®) for Application as Process Scale Biocatalysts

Cross-linked enzyme crystals (CLECs®) are a novel form of immobilised biocatalyst designed for application in large-scale biotransformation processes. In this work we review the production and characterisation of CLECs® prepared from three enzymes (yeast alcohol dehydrogenase I (YADHI), Candida rugosa lipase and α-chymotrypsin) over a range of crystallisation and cross-linking conditions. Optimisation and control of the crystallisation process, with respect to crystal form and enzyme activity yield, was facilitated by the use of triangular crystallisation diagrams which allowed three parameters (e.g. protein concentration, precipitant concentration and pH) to be varied simultaneously. These diagrams showed regions, or 'crystallisation windows', in which particular crystal forms or optimal activity recoveries (up to 87%) could be obtained. They also identified conditions for reproducible scale-up of the lipase crystallisation from 0.5 to 500 mL scale.

In order to evaluate the suitability of a particular batch of CLECs® for large-scale use, a hierarchy of standard tests is proposed. This is designed to expose key properties of the CLECs® relative to each other, and the free enzyme, and to minimise the number of experiments necessary to evaluate each batch of biocatalyst. In general, the CLECs® of each enzyme were found to be more resistant to harsh environmental conditions, such as extremes of temperature and pH and the presence of solvents or proteases, than the free enzymes. Cross-linking of the crystals with glutaraldehyde also yielded mechanically robust catalysts that could withstand the various forces associated with shear in agitated vessels and particle compression in repeated dead-end filtration cycles. The hierarchy of tests proposed here clearly indicated that many of the above properties were also dependent on both the crystal form and size, and the concentration of cross-linking reagent used. Accurate control of the crystallisation conditions used for CLEC® production is therefore vital as this will influence the suitability of the CLECs® for their end use.     

Combined action of Escherichia coli glycogen synthase and branching enzyme in the so-called "unprimed" polyglucoside synthesis.

Mutants of Escherichia coli which are unable to synthesize glycogen were used to study the so-called “unprimed” synthesis of glycogen. The glycogen synthase has been partially purified from these mutants. During the purification, attempts were made to separate the activity which requires the addition of an exogenous primer (primed activity) from the activity which does not require a primer but is highly dependent on the presence of some salts such as citrate and EDTA (unprimed activity). No separation between these two activities could be achieved but the results obtained by chromatography on DEAE-Sephadex indicate that there is a single form of glycogen synthase which is responsible for both unprimed and primed activity. The evidence that a single protein was necessary to catalyze these two reactions was given by the findings that mutants defective in glycogen synthase activity were unable to catalyze glucosyl transfer without added primer. At low concentration, the glycogen synthase purified from a branching enzyme negative mutant catalyzed the unprimed reaction at a slow rate even in presence of salts. A protein activator of this reaction was found in mutants lacking glycogen synthase but not in mutants lacking branching enzyme. The hypothesis that this activator is the branching enzyme itself was supported by the observation that it co-purified with the branching enzyme from a E. coli strain defective in glycogen synthase activity. EDTA or Triton X-100 increased the stimulation of the unprimed synthesis by the branching enzyme. The apparent affinity of the glycogen synthase for glycogen was increased twofold in the presence of EDTA but the branching enzyme further increased the effect of EDTA. The combined action of the glycogen synthase and the branching enzyme on the endogenous glucan associated with the synthase may account for the unprimed activity observed in vitro.     

The role of hydrogen bonding in the cuticular wax of Hordeum vulgare L.

The role of hydrogen bonding in the cuticular wax of Hordeum vulgare L. has been investigated by comparing differential scanning calorimetry and X-ray powder diffraction results of the wax with those of n-alkane mixtures with chain-length distributions resembling that of the wax. It is concluded that hydrogen bonding prevents separation of the short and long chain-length distributions and results in the formation of an amorphous component which is large compared with that of a typical paraffinic wax. It seems that the longer ester chains (39 ≥n≥ 50) bridge the amorphous zone containing chain-ends between two adjacent layers of shorter chains (20 ≥n≥ 33), where n is the number of carbon atoms per chain. In contrast to a paraffinic wax, which has a monolayered structure, this plant wax has a bilayered structure.     

Structural and Biochemical Analysis of a Single Amino-Acid Mutant of WzzBSF That Alters Lipopolysaccharide O-Antigen Chain Length in Shigella flexneri

Lipopolysaccharide (LPS), a surface polymer of Gram-negative bacteria, helps bacteria survive in different environments and acts as a virulence determinant of host infection. The O-antigen (Oag) component of LPS exhibits a modal chain-length distribution that is controlled by polysaccharide co-polymerases (PCPs). The molecular basis of the regulation of Oag chain-lengths remains unclear, despite extensive mutagenesis and structural studies of PCPs from Escherichia coli and Shigella. Here, we identified a single mutation (A107P) of the Shigella flexneri WzzB_SF, by a random mutagenesis approach, that causes a shortened Oag chain-length distribution in bacteria. We determined the crystal structures of the periplasmic domains of wild-type WzzB_SF and the A107P mutant. Both structures form a highly similar open trimeric assembly in the crystals, and show a similar tendency to self-associate in solution. Binding studies by bio-layer interferometry reveal cooperative binding of very short (VS)-core-plus-O-antigen polysaccharide (COPS) to the periplasmic domains of both proteins, but with decreased affinity for the A107P mutant. Our studies reveal that subtle and localized structural differences in PCPs can have dramatic effects on LPS chain-length distribution in bacteria, for example by altering the affinity for the substrate, which supports the role of the structure of the growing Oag polymer in this process.     

Hormonal regulation of glycogen synthase: Insulin decreases protein kinase sensitivity to cyclic AMP

Rat hemidiaphragms incubated with epinephrine exhibited increases in cyclic AMP content and protein kinase activity which were proportional to the logarithm of the hormone concentration from 0.1–2 μM. The fraction of glycogen synthase made independent of glucose-6-P for activity (%I) decreased concomitantly, but correlated only with epinephrine concentrations up to 0.2 μM. Insulin (0–100 mU/ml) increased glycogen synthase %I in a dose-dependent manner with no change in cyclic AMP concentration. Protein kinase activity increased slightly at the lowest insulin concentration, then decreased slightly as glycogen synthase %I increased. Insulin was without effect when administered with a supramaximal dose of epinephrine. In the presence of submaximal epinephrine, insulin produced a dose-dependent increase in glycogen synthase %I which correlated with a decrease in protein kinase activity, without changing cyclic AMP. Insulin had no effect on the increases in cyclic AMP produced by varying levels of epinephrine. However, the activation of protein kinase activity by endogenous cyclic AMP was inhibited in the presence of insulin. The glycogen synthase %I response to epinephrine also was less sensitive in the presence of insulin. Insulin antagonizes the activation of cyclic AMP-dependent protein kinase by epinephrine without altering cyclic AMP levels.     

STUDIES OF NATIVE GLYCOGEN ISOLATED FROM SYNCHRONIZED TETRAHYMENA PYRIFORMIS (HSM)

Native glycogen was isolated from Tetrahymena pyriformis (HSM) by isopycnic centrifugation in cesium chloride density gradients. A density of 1.62 to 1.65 was isopycnic for glycogen. Most of the banded glycogen existed as 35 to 40 mµ particles which had a sedimentation coefficient of 214. These particles were composed of aggregates of 2 to 3 mµ spherical particles. Extraction of glycogen with hot alkali reduced the sedimentation coefficient of native glycogen from 214 to 64.7 and the particle diameter from approximately 40 to 20 mµ and smaller. Cell division was synchronized by a repetitive 12-hour temperature cycle, and glycogen was measured at several times during the cell cycle. The temperature cycle consisted of 9.5 hours at 12°C and 2.5 hours at 27°C. Approximately 90 per cent of the cells divided during the last 1.5 hours of the warm period. The carbohydrate/protein ratio of cells at the end of the cold period was 0.27 and was reduced slightly during the warm period. Glucose was incorporated into glycogen during both periods, although the rate of incorporation was greater during the warm period. No preferential incorporation on the basis of particle size was noted. Incorporation was measured in both native glycogen and KOH-extracted glycogen. Tetrahymena glycogen is compared with rat liver glycogen previously isolated by similar procedures, and the significance of using combined rate-zonal and isopycnic centrifugation for isolating native glycogen is discussed.     

De novo synthesis of Escherichia coli glycogen is due to primer associated with glycogen synthase and activation by branching enzyme.

Previous reports have demonstrated the incorporation of glucose from ADP-glucose into methanol-insoluble and TCA-insoluble fractions in cell extracts of Escherichia coli in the absence of added primer α-glucan. This activity is reduced 6- to 76-fold in cell extracts of three independently isolated glycogen synthase-deficient mutants of E. coli B. Homogeneous preparations of E. coli B glycogen synthase catalyze incorporation of glucose into both methanol- and TCA-insoluble fractions in the absence of added primer. Since glycogen synthase catalyzes these reactions, it is not necessary to propose a protein acceptor glucose or a unique ADP-glucose-glycosyl transferase to catalyze formation of the glucoprotein in E. coli cell extracts to explain glucose incorporation into TCA-insoluble material (R. Barengo et al. (1975) FEBS Lett.53, 274–278). The incorporation of glucose into methanol-and TCA-insoluble fractions is stimulated by 0.25 m citrate and by branching enzyme. Citrate reduces the K_m for the primer, glycogen, about 11- to 15-fold. Branching enzyme can also reduce the concentration of primer required for incorporation of glucose into methanol-insoluble material. The simultaneous presence of both 0.25 m citrate and branching enzyme enables the glycogen synthase reaction rate to proceed at 30% the maximal velocity at a primer concentration of 1 μg/ml. Incorporation of glucose into methanol- or TCA-insoluble material in the absence of primer is completely inhibited by adding α-amylase. Furthermore, incorporation into methanol- or TCA-insoluble material is reduced 13- to 16-fold relative to the reaction occurring in the presence of primer when glycogen synthase is pretreated with glucoamylase and α-amylase. Previous results show that homogeneous preparations of glycogen synthase contain glucan. Heat-denatured glucogen synthase can act as a primer for the glycogen phosphorylase and glycogen synthase reactions. Both the TCA- and methanol-insoluble products form I₂-glucan complexes with wavelength maxima of about 580–590 nm and 610–615 nm, respectively, suggesting that they are mainly linear chain glucans. The products are completely solubilized with α-amylase. The TCA-insoluble product is not solubilized by pronase treatment. The above results strongly suggest that previous reports on formation of glucoprotein primer for glycogen synthesis or on de novo glycogen synthesis in various similar systems is due to endogenous glucan associated with glycogen synthase rather than formation of glucoprotein which then acts as primer for glycogen synthesis.     

Effects of glucose oligomers on the glycogen levels of isolated hepatocytes

Isolated rat hepatocytes appear able to utilize glucose oligomers of the maltose series for the synthesis of glycogen. This utilization is expressed as a stabilization of the glycogen levels during incubation, and also as an incorporation of 14C into glycogen from 14C-labeled glucose polymers. The extent of utilization is only a fraction of that observed with glucose, and it does not appear to be affected by insulin. No extracellular hydrolysis of the polymers seems to be required prior to their uptake by the isolated hepatocytes.