Some properties of β-1,3-glucan hydrolyzing enzymes from the rotifer Brachionus plicatilis

We examined some characteristics of hydrolyticenzymes, especially -1,3-glucanase, to obtain theinformation of cell wall lytic enzymes forrotifers.Crude enzyme (ammonium sulfate fraction) of rotifershydrolyzed starch, -1,3-glucan, glycol chitinand CM-cellulose. Optimum pH for hydrolysis ofstarch and CM-cellulose was 6.5, and that for -1,3glucan and glycol chitin was pH 6.0. Pectic acid,xylan and agarose were not hydrolyzed at pH 3–10.-1,3 glucanase was purified about 73-fold from crudeenzyme by ion-exchange chromatography and gelfiltration. Optimum pH and temperature of the enzymewere 6 and 60 °C, respectively. The molecular weight ofthe enzyme was estimated about 260 kDa by gelfiltration. The enzyme was inhibited byHgCl₂ and MnCl_{_2}.     

Active Site Mutations in Yeast Protein Disulfide Isomerase Cause Dithiothreitol Sensitivity and a Reduced Rate of Protein Folding in the Endoplasmic Reticulum

Aspects of protein disulfide isomerase (PDI) function have been studied in yeast in vivo. PDI contains two thioredoxin-like domains, a and a′, each of which contains an active-site CXXC motif. The relative importance of the two domains was analyzed by rendering each one inactive by mutation to SGAS. Such mutations had no significant effect on growth. The domains however, were not equivalent since the rate of folding of carboxypeptidase Y (CPY) in vivo was reduced by inactivation of the a domain but not the a′ domain. To investigate the relevance of PDI redox potential, the G and H positions of each CGHC active site were randomly mutagenized. The resulting mutant PDIs were ranked by their growth phenotype on medium containing increasing concentrations of DTT. The rate of CPY folding in the mutants showed the same ranking as the DTT sensitivity, suggesting that the oxidative power of PDI is an important factor in folding in vivo. Mutants with a PDI that cannot perform oxidation reactions on its own (CGHS) had a strongly reduced growth rate. The growth rates, however, did not correlate with CPY folding, suggesting that the protein(s) required for optimal growth are dependent on PDI for oxidation. pdi1-deleted strains overexpressing the yeast PDI homologue EUG1 are viable. Exchanging the wild-type Eug1p C(L/I)HS active site sequences for C(L/I)HC increased the growth rate significantly, however, further highlighting the importance of the oxidizing function for optimal growth.     

Comparative study of blocking effects of various retinoids on the occurrence of permanent proliferation of vaginal epithelium in mice treated neonatally with estrogen

Neonatal injections of 20 micrograms 17 beta-estradiol (E2) induced persistent proliferation and cornification of the vaginal epithelium in adult ovariectomized C57 Black/Tw mice. However, permanent vaginal changes were prevented by various retinoids when, simultaneously with E2 treatment, the animals were given injections of 100 micrograms daily dose of retinol, retinol acetate, retinal or of 200 micrograms daily dose of retinol palmitate (RoP). Neonatal injections of a 100 micrograms daily dose of RoP had no preventive effect on the occurrence of E2-induced permanent vaginal changes. This finding suggests that the preventive effect of RoP is weaker than that of other retinoids showing approximately the same degree of prevention. Combined treatment with E2 plus retinoic acid (even a small dose of 20 micrograms) had such a toxic effect on newborn mice that they died within 7 days after birth, while the animals given neonatal injections of 20 micrograms retinoic acid alone survived until the termination of the experiment.     

Localization of triphosphoinositide in the cochlea

Summary Triphosphoinositide (TPI) has been demonstrated to be a receptor for aminoglycosides in the cochlea and may regulate ionic permeability by its binding with Ca⁺⁺. This phospholipid was localized by a protein A-gold technique in the cochlea at the electronmicroscopic level. TPI was prepared by a neomycin column and antibodies to it were raised in rabbits. The antibody used in this study reacted virtually only to TPI among the tested lipids. TPI was localized mainly at stereocilia, cuticular plates, head plates of Deiter's cells, plasma membrane, and mitochondria of various cells in the organ of Corti. In the vascular stria, TPI was found mainly at the plasma membrane of basal infoldings of the marginal cells. Possible physiological and pathophysiological roles of TPI in the cochlea are briefly discussed.     

Survival and transfer in the human gut of poorly mobilizable (pBR322) and of transferable plasmids from the same carrier E. coli

We examined the survival of a host Escherichia coli K-12 bacterium containing two transferable plasmids (pLM2, pSL222-4) and one poorly mobilizable plasmid (pBR322), and the transfer of these three plasmids to endogenous bacteria in the human intestinal tract. The survival of this plasmid-carrying host organism in four human volunteers was 3.5 to 6 days at recovery rates of 10^?1 to 10^?4. This finding was similar to our previous survival data on the same organism bearing a single plasmid. The K-12 strain appeared to be under a strong selective disadvantage in the human gut, since, even when bearing a tetracycline-resistant plasmid, its titer did not increase despite the administration of tetracycline. Studies of transferability showed that, while the transfer-depressed incFII plasmid pSL222-4 transferred at a frequency of 10^?1 in culture, its transfer in the human gut was much less frequent. The number of new recipients per donor cell ingested was about 10^?5, which included new recipients arising by multiplication. The recovery of pSL222-4 transcipients was enhanced by the administration of tetracycline on day 6. Neither the transfer-repressed, broad host range incP plasmid pLM2, nor the plasmid pBR322, could be detected in any endogenous host bacteria. Using the transfer and mobilization frequencies obtained in culture and the number of new recipients of pSL222-4 in the intestinal tract, we estimated that any in vivo mobilization of pBR322 to a new recipient could not occur at a frequency higher than 10^?12.     

Kinetics of the polymerization reaction of tobacco mosaic virus protein: transient-saturation type polymerization reaction.

The kinetics of the endothermic polymerization reaction of tobacco mosaic virus protein in the mild acid region was studied by means of temperature-jump (rising time of 6 sec)-turbidimetry, electron microscopy, and computer simulation. The time course profile of the turbidity increase changed from a normal one to an anomalous one as the size of the temperature-jump was made greater. The anomalous type polymerization profile, which we named the "transient-saturation" type, could be characterized by a rapid increase of turbidity and its transient saturation, and a slow increase to the final level. At a higher concentration of the protein, this transient-saturation effect was more marked, whereas the slow turbidity in the second phase occurred with a higher rate. This transient-saturation type polymerization profile was observed also in a pH-induced polymerization reaction. It was not observed in the case of the N-bromosuccinimide modified tobacco mosaic virus protein under a similar environmental change. By an electron microscopic study and computer simulation, it was revealed that in the first phase, a large number of short polymers were formed, and the concentration of the polymerizing units was rapidly reduced to the equilibrium value, and the polymerization reaction stopped transiently. In the second phase, polymer-polymer associations took place slowly and longer polymers were formed. The revlevance of the present study to the polymerization reaction of actin, myosin, and to a transient-overshoot type polymerization are discussed.     

Hückel framework study on the superconducting vibronic interaction in (AB) N chains

Superconducting vibronic interaction in the vibronic superconductivity motif has been studied in the Hückel framework for (AB) _N chain systems. Within the on-site and nearest-neighbor approximation a new vibronic constant, /L, has been introduced, of which the importance has been discussed. The effect of the vibronic operator, , has also been studied. It is also concluded that the size-dependence of the superconducting vibronic interaction also exists in the (AB) _N chain systems.On leave from the Changchun Institute of Applied Chemistry, Academia Sinica, Changchun, P. R. China, as an STA fellowship awardee hosted by the Institute of Physical and Chemical Research of Japan.     

Cerebro-costo-mandibular syndrome

Summary A 7-month-old boy with the cerebro-costomandibular syndrome is presented. This is the first case report in an Oriental population.15 reported cases in the literature are reviewed.     

Structural study of the sugar chains of alpha-amylases produced ectopically in tumors

About thirty percent of two alpha-amylases produced from a serous papillary cystadenocarcinoma of the ovarium (case 1) and a bronchioloalveolar adenocarcinoma of the lung (case 2) was glycoproteins containing 1 mol of asparagine-linked sugar chain, respectively. The structures of the sugar moieties were found by sequential enzymatic degradation and methylation analysis to be as follows: [(Gal beta 1 leads to 4)0 or 1GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(3)][GlcNAc beta 1 leads to 2Man alpha 1 leads to 3(6)]Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAc and [(Gal beta 1 leads to 4)0 or 1GlcNAc beta 1 leads to 2Man alpha 1 leads to 6][NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3]Man beta 1 leads to 4GlcNAc beta 1 leads to 4(Fuc alpha 1 leads to 6)GlcNAc. Structures of asparagine-linked sugar chains were the same in the tumors of cases 1 and 2 and were incomplete in comparison with those of the parotid amylase.