Effects of sorbitol addition on the action of free and immobilized hydrolytic enzymes in organic media

The effect of the addition of sorbitol on the activity and stability of enzymes was examined by monitoring transesterification reactions performed in organic media at various water activities (a(w) = 0.08 to 0.97). Lipases from Chromobacterium viscosum and Candida rugosa immobilized on celite, and chymotrypsin, free or immobilized on celite, were used. When the sorbitol-containing enzymes were employed, higher reaction rates and less hydrolysis were observed. Immobilization of chymotrypsin resulted in high activity and operational stability, while the nonimmobilized enzyme was stable only in the presence of sorbitol. The activity of all preparations diminished after washing them with pyridine to remove sorbitol. Furthermore, severe stability problems occurred in the preparations lacking sorbitol. Sorbitol treatment, even after removal of the sorbitol itself, improved the activity of nonimmobilized chymotrypsin relative to the washed control. On the other hand, washing to remove sorbitol had a negative effect on the activity of both coimmobilized lipase and coimmobilized chymotrypsin. Addition of a substrate analogue, N-acetyl-L-phenylalanine, to chymotrypsin yielded a preparation that exhibited higher activity than both the control and its sorbitol-containing counterpart. Differential scanning calorimetry measurements revealed that the chymotrypsin-sorbitol complex was stable against thermal denaturation, undergoing transition at a high temperature (89 degrees C). The transition temperatures of the substrate-containing chymotrypsin and of the control were identical (72 degrees C). (c) 1995 John Wiley & Sons, Inc.     

Origin of 33 kDa protein of vitelline membrane of quail egg: Immunological studies

The inner layer of vitelline membrane is an investment of avian ovum at the time of ovulation, but its formation is poorly understood. In order to elucidate the origin of the inner layer of vitelline membrane, a 33 kDa protein, one of the components of the inner layer, was purified from quail eggs and polyclonal antibody was raised against this protein. The tissue distribution of protein interacted with the antibody was studied by Western blotting technique. No immunoreactive component could be observed in extracts of liver, kidney, heart, lung, small intestine, brain, infundibulum, albumen-secreting region of oviduct, uterus, and wall of small white follicles. The intensive band was detected in the granulosa layer, which was isolated from the large preovulatory follicles as a monolayer of granulosa cells sandwiched between the inner layer of vitelline membrane and the basal lamina. The granulosa cells isolated from the granulosa layer also reacted with this antibody. Theca layer had no immunoreactive components. The position of the band of the 33 kDa protein on SDS-PAGE was sifted to higher molecular weight in follicular tissues as compared with that in the laid eggs, indicating that the structural change of the protein occurs after ovulation. These studies indicate that the material reactive to the antibody raised against a 33 kDa protein of quail vitelline membrane is synthesized by the granulosa cells.     

A scanning electron microscope study of Craspedella sp. from the branchial chamber of redclaw crayfish,Cherax quadricarinatus,from Queensland,Australia

Epidermal topography was examined, including papillate ridges, grooves and ciliated sensory papillae of Craspedella sp. from the branchial chamber of redclaw crayfish, Cherax quadricarinatus, from Queensland, Australia. Rhandites were observed to discharge from ducts opening mainly in a small distal region of the ventral epidermis of the three central (of five) tentacles. These regions, devoid of ciliated sensory papillae, serve to adhere the anterior end of the worms during locomotion. Secretions from glands associated with the posterior attachment organ were observed to discharge from pores on the outside region of the ventral surface of the disc.A comparison of various scanning electron microscopy (SEM) fixation techniques showed that (1) hot fixatives at 90 °C provide most information on the largest number of epidermal structures and (2) different fixation regimes highlight different epidermal features.     

International study on Artemia. LIV. Morphological study of Artemia with emphasis to Old World strains. II. Parthenogenetic populations

Eleven morphometric and one meristic character in 15 parthenogeneticArtemia populations have been studied by using discriminant andcluster analysis as well as scanning electron microscopy.Discriminant analysis revealed five main groups of morphologicalpatterns: (i) the coastal Chinese populations together with apopulation from Kazakhstan, (ii) the inland Chinese salt lakepopulations, (iii) the Greek populations, (iv) one African populationfrom Namibia and (v) a Chinese population from Xuyu (Jiangsuprovince). Cluster analysis was not always in agreement withdiscriminant analysis and these results are discussed.     

The Structure and Characterization of the Surface Protein Layer of a Comamonas-Like Organism

The regular surface layer of a strain of a Comamonas-like organism was examined by electron microscopy. The surface layer protein was easily extracted from the cell surface by a 2.5 M solution of lithium chloride. The protein subunit has a molecular size of 32,000 daltons, but usually forms a large aggregate of more than 1,200,000 daltons. In the extract it formed a regular array of p4 symmetry and was observed to be intimately associated with fragments of lipopolysaccharide. The size of a subunit determined by the negative staining method and the image processing method measured 5.2 × 6.4 nm (width and length), was arranged in a cobblestone-like pattern, and was located in a lattice space measuring 13.0 nm square.     

Effects of butylated hydroxyanisole,butylated hydroxytoluene and tertiary butylhydroquinone on growth and luteoskyrin production byPenicillium islandicum

Butylated hydroxyanisole (BHA), Butylated hydroxytoluene (BHT) and tertiary butylhydroquinone (TBHQ) alone in cultural media were tested for the inhibition of growth and luteoskyrin production by two toxigenic strains ofPenicillium islandicum UST-11 andP. islandicum HLT-6. In potato dextrose agar, the concentrations of BHA and TBHQ from 0.2 mg/disc, BHT from 5.0 mg/disc did affect the growth of both tested strains, but the initial concentrations of these antioxidants to reduced luteoskyrin production by UST-11 strain were BHA 0.5 mg/disc, BHT 1.0 mg/disc and TBHQ 0.4 mg/disc, while for HLT-6, BHA 0.4 mg/disc, BHT and TBHQ were 0.2 mg/disc, respectively. In grainy and powdery rice media, the effects of BHA, BHT and TBHQ on luteoskyrin production byP. islandicum UST-11 and HLT-6 were clearly demonstrated. The efficiency of the inhibitory effect was not only closely related to the concentration of antioxidants, but also completely inhibited the luteoskyrin production at a concentration of 200 mg/kg or higher. Also, the antioxidants at a concentration higher than 20 mg/kg reduced significantly the growth and luteoskyrin production by both strains ofP. islandicum.     

Structural energetics of barstar studied by differential scanning microcalorimetry.

The energetics of barstar denaturation have been studied by CD and scanning microcalorimetry in an extended range of pH and salt concentration. It was shown that, upon increasing temperature, barstar undergoes a transition to the denatured state that is well approximated by a two-state transition in solutions of high ionic strength. This transition is accompanied by significant heat absorption and an increase in heat capacity. The denaturational heat capacity increment at approximately 75 degrees C was found to be 5.6 +/- 0.3 kJ K-1 mol-1. In all cases, the value of the measured enthalpy of denaturation was notably lower than those observed for other small globular proteins. In order to explain this observation, the relative contributions of hydration and the disruption of internal interactions to the total enthalpy and entropy of unfolding were calculated. The enthalpy and entropy of hydration were found to be in good agreement with those calculated for other proteins, but the enthalpy and entropy of breaking internal interactions were found to be among the lowest for all globular proteins that have been studied. Additionally, the partial specific heat capacity of barstar in the native state was found to be 0.37 +/- 0.03 cal K-1 g-1, which is higher than what is observed for most globular proteins and suggests significant flexibility in the native state. It is known from structural data that barstar undergoes a conformational change upon binding to its natural substrate barnase.(ABSTRACT TRUNCATED AT 250 WORDS)     

A thin quartz cell suitable for vacuum ultraviolet absorption and circular dichroism measurements

The design of a thin quartz cell suitable for absorption and circular dichroism measurements in the vacuum ultraviolet is described. Important features of the cell are (1) that it can be disassembled for cleaning and reproducibly reassembled with path lengths up to 0.3 mm, and (2) that strain in the windows from the compressed sample can be relieved by a sample overflow port. The latter feature allows the cell to be used for circular dichroism as well as absorption measurements.     

Vascular morphology of the bovine spermatic cord and testis

Summary The highly coiled testicular artery within the bovine spermatic cord has a constant luminal diameter but a continuously decreasing mural thickness. The pampini form plexus is composed of three interconnected venous networks differing in mesh sizes and calibres. The large veins of the first network display pouches and permanent constrictions, which may serve as throttle devices. The constitutents of the third network are venules or venous capillaries with diameters between 10 and 20 m; they favor a periarterial position or even occupy the media-adventitia border of the testicular artery. All plexus veins are devoid of valves. The existence of true arteriovenous anastomoses between smaller branches of the testicular artery and plexus veins was established by serial sections. The vascular morphology of the spermatic cord is discussed with special attention to a postulated venous-arterial steroid transfer in this region.Supported by the Deutsche Forschungsgemeinschaft and the Stiftung zur Förderung der wissenschaftlichen Forschung an der Universität Bern     

The permeability coefficient of the wall of a villous membrane

Summary The equations hitherto used to correct the permeability coefficient for the unstirred layer influence are valid only for flat membranes. Therefore, appropriate equations for membranes with a villous surface (e.g., small intestine) have been derived. They take into account the non-linear concentration gradient in the intervillous part of the unstirred layer. Quantitative information about the geometry of the villous surface and the unstirred layer thickness are needed to calculate the permeability coefficient of the membrane wall (e.g., intestinal epithelium). The concentration of highly permeable substances drops sharply already in the upper part of the intervillous space, so that the tips of the villi function as effective absorbing area. The intervillous concentration gradient of a substance with a low permeability coefficient is so small, that such a substance is absorbed by the total surface area of the villous membrane. The effective absorbing area of substances with intermediate permeability coefficient lies between the described limits.