Functionally important lysine residues in inorganic pyrophosphatase from E. coli. II. Isolation and characteristics of modified tryptic peptide and analysis of the functional role of lysine residues controlling enzyme activity

Inactivation of inorganic pyrophosphatase from E. coli by pyridoxal-5'-phosphate is due to the modification of a lysine residue located in the tryptic peptide with the Asp-Leu-Pro-Glu N-terminal sequence. In course of the enzymatic process this lysine-residue appears to be in the protonated state and either operators as the proton donor for the product of the enzymatic reaction or is involved in stabilization of the transition state.     

Structure of a protein catalyzing the formation of 11 cis-retinal in the visual cycle of invertebrate eyes

A pigment made up of a protein able to bind retinal as well as retinol is described. The molecule consists of a dimer with a molecular weight of 50,000 which binds one molecule of retinal. The binding site for retinal is a Schiff base buried in the interior of the protein. Retinol is probably bound to the protein in the same site as for retinal, although not covalently, as suggested by the absorbance spectra. The protein, extracted from honeybee retina, is involved in visual pigment metabolism, and its structure may elucidate the mechanism of the stereospecific photoisomerization of all trans-retinal to 11-cis-retinal.     

Plant regeneration from protoplasts of the monocotyledonous Haworthia magnifica v. Poelln.

Calli were initiated from flower buds, gynoecia and inflorescence segments of Haworthia magnifica v. Poelln. and subcultured on solid medium. Two liquid culture steps were necessary to prepare the calli for the isolation of protoplasts capable of sustained cell divisions. Plants were regenerated from protoplast-derived calli. The influence of both the osmolality of the culture media and exudates on the viability of protoplasts and protoplast-derived cell colonies is briefly discussed.     

Characterization of a light-dependent glutamate synthase activity in Chlamydomonas reinhardtii

Photosynthetically active vesicles prepared from Chlamydomonas reinhardtii retained a light-dependent glutamate synthase activity which was highly specific for 2-oxoglutarate (K_m=2.1 mM) and L-glutamine (K_m=0.9 mM) as amido group acceptor and donor respectively. This activity was inhibited by azaserine, p-hydroxymercuribenzoate and 3-(p-chlorophenyl)-1,1-dimethyl urea.Light-dependent synthesis of glutamate was also obtained by coupling Chlamydomonas photosynthetic particles to purified ferredoxin-glutamate synthase, using ascorbate and 2,6-dichlorophenol-indophenol as electron donor. This system was also specific for 2-oxoglutarate (K_m=1 mM) and L-glutamine (K_m=0.8 mM) as substrates, and was stimulated by dithioerythritol. Azaserine and p-hydroxymercuribenzoate, but not 3-(p-chlorophenyl)-1,1-dimethyl urea, inhibited the reconstituted activity; high concentrations of 2-oxoglutarate were inhibitory.Abbreviations A Absorbance - CCP p-Trichlorometoxi-carbonylcyanide-phenylhydrazone - Chl Chlorophyll - CMU 3-(p-Chlorophenyl)-1,1-dimethyl urea - DPIP 2,6-Dichlorophenol-indophenol - DTE Dithioerythritol - MSX L-Methionine, D-L, sulfoximine - MV Methyl viologen     

Pheophytin-mediated energy storage of photosystem II particles detected by photoacoustic spectroscopy

The photoacoustic (PA) characteristics (energy storage and heat dissipation) of photosystem II (PSII) core-enriched particles from barley were studied (i) in conditions where there was electron flow, i.e., in the presence of a combination of the electron acceptor K₃ Fe (CN)₆, referred to as FeCN, and the electron donor diphenylcarbazide (DPC), and (ii) in conditions where electron flow was suppressed, i.e., in the absence of FeCN and DPC. The experimental data show that a decrease of heat dissipation with a minimum at 540 nm can be interpreted as energy storage resulting from the presence of pheophytin (Pheo) in the PSII particles. On account of the capability of the PA method to measure the energy absorbed by the chromophores which is converted to heat, it is suggested that the PA detection of Pheo present in the PSII complex will permit to clarify the function of processes involving non-radiative relaxation of excited states in P680-Pheo-Q_A interactions.Abbreviations -Car -Carotene - Chl Chlorophyll - DPC Diphenylcarbazide - EPR Electron Paramagnetic Resonance - FeCN potassium ferricyanide - HEPES N-2-hydroxyethylenepiperazine-N-2-ethanesulfonate - P680 reaction center of PSII - PA Photoacoustic - Pheo pheophytin - PSI photosystem I - PSII photosystem II - Q_A primary electron acceptor of PSII     

Transforming potential of a myc-containing variant of feline leukemia virus in vitro in early-passage feline cells.

We studied a naturally occurring variant of feline leukemia virus (FeLV) in which the oncogene myc has substituted for a portion of the viral structural genes (myc-FeLV). myc-FeLV was rescued by replication in the presence of FeLV as helper, and its biological activity was examined in early-passage feline cells in vitro. Infection of leukocytes from peripheral blood, spleen, or thymus, or of kitten fibroblasts did not immortalize these cells or alter them morphologically. Northern blot (RNA blot) analysis of virion RNA prepared from the supernatant of infected cells demonstrated the 8.2-kilobase genome of FeLV, but did not demonstrate the 5.0-kilobase genome of myc-FeLV. Apparently, the myc-FeLV genome was lost in the absence of the selective pressure of transformation. In contrast, infection of embryonic fibroblasts with myc-FeLV(FeLV) rendered these cells capable of greatly increased, if not infinite, proliferative potential. The cells were morphologically altered compared with controls and were only loosely adherent to the substrate. The cells failed to proliferate in semisolid medium and did not form tumors when inoculated subcutaneously into athymic mice. Blot analyses demonstrated the presence and expression of integrated proviral DNAs of both FeLV and myc-FeLV in these cells. They appear, then, to represent cells partially transformed by infection with myc-FeLV(FeLV). The action of feline v-myc in early-passage cells in vitro was compared to that of avian v-myc.     

3'-5' cyclic-guanosine monophosphate increase in rat brain hippocampus after gamma-hydroxybutyrate administration. Prevention by valproate and naloxone

An increase (123%) of cyclic GMP (cGMP) was observed in the hippocampus of the rat killed by microwave irradiation 45 min after administration of 500 mg/kg gamma-hydroxybutyrate (GHB) IP. This increase is time and dose dependent. No modification in cyclic nucleotide content was observed in striatum and in cerebellum. As the role of GHB has been implicated in neurotransmission, the fact that this compound increases cyclic GMP accumulation in hippocampus in vivo may represent a mechanism by which the actions of GHB are mediated at the cellular level. Valproate (400 mg/kg) or naloxone (10 mg/kg) pretreatment completely abolish the cGMP increase due to GHB. A GABAergic and/or opiate phenomenon may be involved in the mechanism of GHB induced increase of cGMP.     

The association between spontaneous pyelonephritis and maturity-onset diabetes mellitus in male MM mice

Blood glucose and glucose tolerance tests demonstrated that many male MM mice are diabetic. Serial urine sampling showed that the diabetes occurred only in mature MM males and consisted of a single self-limiting episode. Histological examination of the pancreas, together with measurements of body weight, glycosylated haemoglobin and plasma insulin, revealed that the diabetes was of the maturity-onset insulin-resistant type. Bacteriological examination of the urine samples showed that urinary tract infection, a known feature of male MM mice, occurred in the diabetics but only after the onset of hyperglucosuria. It was concluded that the high urinary glucose levels of diabetic MM males are of prime importance in the aetiology of the renal infection which occurs rarely in non-diabetic MM males or in other strains in the colony. An infectious aetiology for the diabetes per se was excluded by the existence of diabetes in germfree MM males.     

Antitrypanosomal action of cis-diamminedichloroplatinum (II) analogs

The present report deals with the alterations produced by cis-diamminedichloroplatinum (II) (DDP), and 2 of its analogs: cis-Pt(II)(tranylcypromine)2Cl2 and cis-Pt(II)(benzothiazole)2Cl2 in cultured epimastigote forms of Trypanosoma cruzi. Studies have been performed at the ultrastructural level and the inhibitory effect of these complexes on macromolecule synthesis, evaluated by 3H-thymidine, 3H-uridine, and 3H-leucine incorporation, has been investigated. DDP at concentrations of 50 and 100 micrograms/ml does not inhibit significantly the incorporation of radioactive precursors, but a clear decrease was observed with the 2 analogs. Eight hours of treatment at a concentration of 10 micrograms/ml rendered in all 3 cases an increase in autophagic vacuoles and lipids as well as an abnormal condensation of the nucleus chromatin.