Purification and characterization of α-glucosidases produced by Saccharomyces in response to three distinct maltose genes

α-Glucosidases or maltases (EC 3.2.1.20) were purified to electrophoretic homogeneity from a respective strain of Sacchromyces cerevisiae which carries a single MAL gene, either MALα, MALβ or MALγ, using gluconate-Sepharose affinity chromography and isoelectrofocusing. Of these maltases, two types of maltase were obtained from the MALγ strain, the pI values of which were 5.6 and 5.9. From the MALα and MALβ strain was obtained only one type of maltase with the pI at 5.6 which was identical to one of the maltases from the MALγ strain. These four maltases possessed the same properties, except for pI. They were monomers with molecular weights of between 66 000 and 67 000. With regard to the substrate specificity, they hydrolyzed maltose and sucrose exclusively but not α-methulglucoside nor maltooligosaccharide. They did not differ in immunological properties.     

Use of a bioreactor consisting of sequentially aligned L-glutamate dehydrogenase and L-glutamate oxidase for the determination of ammonia by chemiluminescence

A chemiluminometric method for the automated flow injection analysis of ammonia is described. The essence of the invention is the use of a bioreactor consisting of both immobilized L-glutamate dehydrogenase (GLDH) and L-glutamate oxidase (GLXD), which are sequentially aligned in this order in a minicolumn measuring 2.0 X 20 mm. The unidirectional constant flow of liquid through the column reactor minimizes the reversed diffusion of the solutes so that the following sequence of reactions is ensured. Thus, ammonia to be determined is first transformed by GLDH into L-glutamate, which then produces hydrogen peroxide by GLXD. Hydrogen peroxide in the effluent from the column is then determined by its chemiluminescence upon admixing with luminol and potassium ferricyanide. The present method gives linearity of the standard curve for ammonia up to 1.0 mM. It is at least 100 times more sensitive than the conventional method for ammonia assay using ultraviolet absorption measurement.     

Effect of high-intensity endurance training on isokinetic muscle power

The purpose of this study was to determine the effects of high-intensity endurance training on isokinetic muscle power. Six male students majoring in physical-education participated in high intensity endurance training on a cycle ergometer at 90% of maximal oxygen uptake (VO2max) for 7 weeks. The duration of the daily exercise session was set so that the energy expenditure equalled 42 kJ.kg-1 of lean body mass. Peak knee extension power was measured at six different speeds (30 degrees, 60 degrees, 120 degrees, 180 degrees, 240 degrees, and 300 degrees.s-1) with an isokinetic dynamometer. After training, VO2max increased significantly from mean values of 51.2 ml.kg-1.min-1, SD 6.5 to 56.3 ml.kg-1.min-1, SD 5.3 (P less than 0.05). Isokinetic peak power at the lower test speeds (30 degrees, 60 degrees and 120 degrees.s-1) increased significantly (P less than 0.05). However, no significant differences in muscle peak power were found at the faster velocities of 180 degrees, 240 degrees, and 300 degrees.s-1. The percentage improvement was dependent on the initial muscle peak power of each subject and the training stimulus (intensity of cycle ergometer exercise).     

General recombination mechanisms in extracts of meiotic cells

RecA-like proteins have been purified from somatic and meiotic cells of mouse and lily. The rec proteins have been designated s-rec and m-rec to indicate their respective tissues of origin. The two proteins differ in molecular weight and in their response to temperature, the latter being consistent with the optimal temperature for physiological function of their tissues of origin. There is a major increase in m-rec protein with the entry of cells into meiosis, the peak of activity being early pachytene. Extracts of the cells and also those of yeast (Saccharomyces cerevisiae) have been prepared that have the capacity to catalyze homologous recombination. These extracts behave similarly to the m-rec proteins upon entry of cells into meiosis. Yeast transferred to sporulation medium displays a 100-fold increase in the recombination activity of the extract at about the time of entry into meiosis. The occurrence of peak levels of m-rec and recombination activity in extracts from cells in early pachytene points strongly to that stage as the time at which the enzymatic phase of recombination occurs.     

Hormonal regulation of berberine production in cell suspension cultures of Thalictrum minus

Effects of auxin and cytokinin on cell growth and alkaloid production in cell suspension cultures of Thalictrum minus were examined in an attempt to increase the productivity of a medicinal compound, berberine. In Linsmaier and Skoog medium containing auxin such as 2,4-D (1 M), the cultured cells grew rapidly, producing little berberine. On the other hand, the berberine-producing activity was remarkably enhanced by simultaneous administration of auxin and cytokinin, although cell growth was inferior. In particular, for the combination of NAA (60 M) and 6-benzylaminopurine (10 M), the yield of berberine was as high as 20 mg/30 ml medium after 2 weeks of culture. Furthermore, most of the berberine produced by the cells was released into the liquid medium, in which an excess of berberine crystallized. The results of the present experiments are suggestive of an advantage in adopting a two-stage culture method for the production of berberine in fermentor systems.Abbreviations LS Linsmaier and Skoog - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA 1-naphthaleneacetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - BAP 6-benzylaminopurine     

Enzymatic difference between laticifers and cultured cells of papaya

Cell suspension cultures of Carica papaya were found to produce a papain-like enzyme showing an amidase activity similar to papain. Experiments suggested that the enzyme was an SH-enzyme, but an immunological test indicated the absence of papain in cultured cells. The isoelectric point (4.3) of the enzyme of cultured cells was the same as that of the leaf extract, but was different from that of papain or other amidases in the latex of the fruit.     

Induction of berberine biosynthesis by cytokinins in Thalictrum minus cell suspension cultures

Production of berberine could be induced by adding 6-benzylaminopurine (BAP) to Thalictrum minus cells, cultured in suspension in a medium containing 2,4-dichlorophenoxyacetic acid (2,4-D), early in the growth cycle. In the presence of BAP, the precursor, L-tyrosine, was rapidly converted into berberine which was then released into the medium, whereas substantial amounts of the intermediates, tyramine and dopamine, accumulated in non-berberine-producing cells grown in the same 2,4-D-containing medium without BAP. These results suggest that BAP activated enzymatic reactions subsequent to the formation of the amines in the biosynthesis of berberine.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - BAP 6-benzylaminopurine - NAA 1-naphthaleneacetic acid - IAP 6-isopentenylaminopurine - LS medium Linsmaier-Skoog medium - Growth medium LS medium containing 10^-6 M 2,4-D     

Delta-Cytomembranen und lamelläre Cytosomen

Zusammenfassung Im Epithel der Kiemenbüschel von Axolotl (Amblystoma mexicanum) findet sich ein besonderer Typ von cytoplasmatischen Membranen, den wir mit -Cytomembran bezeichnen. -Cytomembranen sind schichtweise in Schleifen oder Spiralen angeordnet und bestehen aus einer 30–45 Å dicken osmiophilen Schicht mit einem 60 Å breiten intermembranösen Intervall. Die -Cytomembranen differenzieren sich im perinucleären Bereich des Cytoplasmas aus einer homogenen, elektronendichten Substanz und stellen die Vorstufen der lamellären Cytosomen dar. Die -Cytomembranen und die lamellären Cytosomen sind aus einem Kohlenhydrat-Protein-Komplex mit möglicher Bindung an Phospholipoide zusammengesetzt. Wir glauben, daß diese besondere Gruppe von submikroskopischen Strukturen wichtige Funktionen für die Synthese der Mucopolysaccharide im allgemeinen und für die Schleimsekretion im speziellen hat.Stipendiat der Deutschen Forschungsgemeinschaft.