Sialic acid moiety is responsible for the charge heterogeneity and the biological potency of rat lutropin

To elucidate a possible role of sialic acid moiety in the electrical heterogeneity of rat pituitary lutropin, seven components separated were individually treated with neuraminidase. Some intermediates with isoelectric points corresponding to the native components were concomitantly seen at the serial stages of the enzyme treatment. All the treated components showed an isoelectric point of about 10.0 which was the same to the isoelectric point of one of the seven components. Desialylation of the components with less biological activity caused enhancement of the in vitro cyclic AMP producing- and testosterone producing-activities as well as the binding activity to the receptor. It is concluded that the number of sialic acid moiety in lutropin is responsible for the charge heterogeneity and the biological potency of the hormone.     

Location of the SH group of the alkali light chain on the myosin head as revealed by electron microscopy

The location of the single cysteinyl residue of the alkali light chain on the myosin head was determined by electron microscopy. The cysteinyl residue of isolated alkali light chain 2 was biotinylated and the light chain was exchanged with that of heavy meromyosin in 4.7 M-NH4Cl. Avidin was attached to the biotin in the heavy meromyosin and the complex was rotary shadowed and observed in the electron microscope. The distance from the head-rod junction to the centre of avidin was 8(+/- 3) nm (mean value +/- standard deviation: n = 105).     

Characteristics of major mutagenicity of instant coffee

Detailed quantitative studies on the mutagenicity of methylglyoxal showed that its contribution to the total mutagenicity of instant coffee on S. typhimurium TA100 was minor although we reported previously (Kasai et al., 1982) that its contribution to the mutagenicity of freshly brewed coffee was about 50%. Cysteine suppressed the mutagenicity of methylglyoxal and of methylglyoxal when added to instant coffee, but did not affect the mutagenicity of coffee itself. Catalase suppressed most of the mutagenicity of coffee, but not that of methylglyoxal or of methylglyoxal added to coffee.     

Is the initiation of macronuclear DNA synthesis in Euplotes dependent on micronuclear functions?

To determine whether the micronucleus makes essential contributions during asexual reproduction, observations were made on cells of Euplotes octocarinatus from which the micronucleus had been removed with a micropipette. Most cells underwent one postenucleation division, then became arrested in macronuclear G1, slowed down in food uptake, developed macronuclear deformations, and finally died. Such cells could be rescued if a micronucleus was reimplanted before macronuclear deformations had developed. When provided with a new micronucleus, cells initiated macronuclear DNA synthesis about 12-16 h later. The data suggest that the micronucleus is involved in the control of the cell's transition from macronuclear G1 to S, and a model is proposed which postulates that in Euplotes macronuclear DNA synthesis is initiated when a micronucleus-encoded "initiator protein" has accumulated to a critical amount.     

Formation of giant protoplasts from protoplasts of Pleurotus cornucopiae by the cell wall lytic enzyme

Summary When purified protoplasts of Pleurotus cornucopiae IFO9614 were incubated with a mixture of cell wall lytic enzymes, they were found to increase their size. Their average diameter increased from 4.3 m to 31 m after 65 h incubation at 24° C. The presence of cellulase ONOZUKARS in the enzyme mixture had a significant effect on the formation of giant protoplasts. Regeneration frequency of giant protoplasts in a medium containing 0.5 M sucrose was 3.5%, approximately six times that of normal protoplasts.     

Electron microscopic studies on myosin molecules from chicken gizzard muscle III. Myosin dimers

Previously, we (Onishi, H. & Wakabayashi, T. (1982) J. Biochem. 92, 871) reported that the ATP-induced disassembly of chicken gizzard "thick filaments" resulted in myosin monomers with "looped" tails. In the present study, we found that these monomers assembled themselves into antiparallel dimers when they were placed in a medium of low ionic strength (approximately 2 mM).     

Proximity of reactive lysyl residue to the antigenic site in rabbit skeletal myosin against the monoclonal antibody (MF-18) generated to chicken skeletal myosin

MF-18, one of the monoclonal antibodies generated to chicken myosin, cross-reacted with rabbit skeletal myosin subfragment-1 (S1). Utilizing an improved procedure of immuno-blotting, a decrease in reactivity of MF-18 to S1 by trinitrophenylation was observed. This indicates that the reactive lysyl residue is very close to the hapten site. This is consistent with the evidence that the hapten site resides in the 26,000 dalton tryptic fragment of S1. Use of such antibodies as labels may open the way to determining the location of specific hapten sites in the three-dimensional image of actin-S1 complex reconstructed from the electron micrographs.     

Reaction mechanism of Ca2+-dependent adenosine triphosphatase of sarcoplasmic reticulum. ATP hydrolysis with CaATP as a substrate and role of divalent cation

ATP hydrolysis with CaATP as a substrate was characterized at 0 degrees C and pH 7.0 using purified ATPase preparations of sarcoplasmic reticulum and compared with that with MgATP as a substrate. The maximal rate of enzyme phosphorylation and the Km value for the phosphorylation were 8 to 10 times less for CaATP than for MgATP. Each substrate appeared to act as a competitive inhibitor with respect to the other in enzyme phosphorylation. The phosphoenzyme formed from CaATP turned over slowly because the conversion rate of the ADP-sensitive (E1P) to ADP-insensitive (E2P) phosphoenzyme was very slow. E2Ps, formed from both CaATP and MgATP, were similar in that KCl, MgCl2, or ATP accelerated their decomposition. Their sensitivity to KCl and/or ATP was retained even after a long incubation with excess EDTA. When the enzyme had been phosphorylated from CaATP, calcium remained bound to the enzyme even in the presence of excess EDTA. The observed parallelism between the amount and behavior of the enzyme-bound calcium and those of E2P strongly suggests that 1 mol of E2P has 1 mol of tightly bound calcium. During steady state ATP hydrolysis with CaATP as a substrate, a significant amount of the enzyme-ATP complex accumulated as a reaction intermediate because of slow dissociation of CaATP from the CaATP-enzyme complex and slow enzyme phosphorylation from the CaATP-enzyme complex. These results indicate that Mg2+ is not essential for the turnover of the calcium pump ATPase. It was proposed that the metal component of the substrate basically determines affinity of the substrate to the enzyme and the catalytic mechanism of subsequent reaction steps.     

The effect of 2-deoxy-D-glucose on insulin release by neonatal rat B cells in culture

The culture for 7 days in medium with 5.5 mM glucose and 1 mM 2-deoxy-D-glucose enhanced the glucose sensitivity of neonatal rat B cells, and even stimulated their growth in vitro. Also, 2-deoxy-D-glucose supplementation maintained insulin release evoked by leucine and 2-ketoisocaproate from B cells at day 7 at levels several times higher than at day 1. The effect of leucine was greatly augmented by glutamine, whereas that of the 2-keto acid remained almost unchanged irrespective of the presence of glutamine. These results suggest an increase in oxidative catabolism of medium nutrients in B cells grown in medium with 2-deoxy-D-glucose for 7 days, and such metabolic changes may promote the growth of B cells in vitro.     

On the structure of biological membranes: The double-tiered pattern

A set of six biological membranes has been examined electron microscopically in positively-stained sections at both low and high resolution. At low resolution, all six membranes exhibit a railroad track pattern, while at high resolution all six membranes exhibit a pattern of a double-tier of globular particles. The correspondence between the railroad track pattern and the double-tiered pattern is considered. The relationship of the double-tiered pattern to the bimodal properties of the constituent protein and phospholipid molecules is discussed.