Relative biological efficiency of neutrons (2 MeV) and X-rays for mastocytoma cell division capacity]

The effects of 2 MeV neutrons were compared with those of 200 kV X-rays for the criterion of lethality in mastocytoma cells of ascites type. The cells prepared as suspension were exposed to the radiations in vitro under aerobic condition and transplanted in the abdominal cavity of mice. Their viability was estimated according to the growing speed of the excreted amount of urinary 5-HIAA in the host animal. The RBE varied between a value of 3.1 at the 10 per cent level of cell survival to 1.8 for the mean lethal dose (Do).     

Sigmoidal stimulation by solutes of light-induced proton translocation in thylakoid membranes

A sigmoidal curve was obtained for the relationship betweenthe stimulation of light-induced proton uptake and the concentrationof salts in a suspending medium for thylakoid membranes. Substitutionof sucrose for the salts also resulted in a sigmoidal curve.It changed into a hyperbolic curve with salts when the mediumalready contained sucrose. The results are discussed in relationto the structural arrangement of the thylakoid membranes bythe osmotic effect of the solutes. (Received February 10, 1975; )     

Analysis of yeast prp20 mutations and functional complementation by the human homologue RCC1, a protein involved in the control of chromosome condensation

Summary A DNA fragment that codes for the 364 amino-terminal amino acid residues of a putative Bacillus subtilis SecA homologue has been cloned using the Escherichia coli SecA gene as a probe. The deduced amino acid sequence showed 58% identity to the aminoterminus of the E. coli SecA protein. A DNA fragment which codes for 275 amino-terminal amino acid residues of the B. subtilis SecA homologue was expressed in E. coli and the corresponding gene product was shown to be recognized by anti-E. coli SecA antibodies. This polypeptide, although only about 30% the size of the E. coli SecA protein, also restored growth of E. coli MM52 (secA ^ts) at the non-permissive temperature and the translocation defect of proOmpA in this mutant was relieved to a substantial extent.     

Distinct mode of G protein activation due to single residue substitution of active IGF-II receptor peptide Arg2410-Lys2423: evidence for stimulation acceptor region other than C-terminus of Gi alpha.

Arg2410-Lys2423 (RVGLVRGEKARKGK, peptide 14) of the human insulin-like growth factor II receptor directly activates Gi and deletion of C-terminal 4 residues from peptide 14 nullifies this activity. A study was thus made of the effects of peptides modified in the C-terminal structure. RVGLVRGEKAAKGK and RVGLVRGEKARKGA scarcely activated Gi, whereas RVGLVRGEKARAGK (peptide A5) activated Gi as much as peptide 14 did. However, peptide A5 action did not depend on Mg2+ concentration and was little affected by pertussis toxin modification of Gi alpha. Peptide A5 may thus recognize the region on Gi alpha that is distinct from the extreme C-terminus. It is consequently considered that (i) the first and the last basic residues in the C-terminal motif of peptide 14 determine the capacity for recognition of Gi and (ii) there is a region different from the C-terminus of Gi alpha, through which the C-terminal second basic residue-altered peptide 14 activates Gi in a Mg(2+)-independent manner.     

Oxidative DNA damage in tissues of English sole (parophrys vetulus) exposed to nitrofurantoin

Juvenile English sole were exposed intramuscularly to nitrofurantoin (NF) and the levels of 8-hydroxy-2′deoxyguanosine (8-OH-dG) in liver, kidney and blood were determined using reversed-phase HPLC with electrochemical detection. Identification and quantitation of the 8-OH-dG in the samples was accomplished by comparison with standard 8-OH-dG, which was characterized by UV spectroscopy and fast-atom bombardment mass spectrometry. The levels of hepatic 8-OH-dG increased (r² = 0.59, P = 0.015) with the dose of NF (0.10 – 10 mg NF/kg fish). In kidney and blood, however, the levels of 8-OH-dG were significantly higher than controls only at the highest dose tested. The level of binding in liver ranged from 0.37 to 0.76 fmol 8-OH-dG/μg DNA. The levels of hepatic 8-OH-dG reached a maximum (approx. 1 fmol 8-OH-dG/μg DNA) between 1 and 3 days after exposure, followed by a decrease to control levels (approx. 0.25 fmol 8-OH-dG/μg DNA) at 5 days post-exposure. These data demonstrate the first direct evidence for the formation of oxidized DNA bases resulting from the metabolism of a nitroaromatic compound by fish.     

Possible requirement of serum progression factors for transformation of BALB/c 3T3 fibroblasts by v-ras p21.

To form colonies in soft agar, ras-transformed 3T3 fibroblasts require serum. We examined what growth factors in serum were essential for ras-induced transformation. Temperature-sensitive (ts) v-Ki-ras-transfected BALB/c 3T3 cells were used to strictly control both the activity of the ras protein and the cell cycle. When G0-arrested ts cells were cultured with 10% serum at a permissive temperature, greater than 50% of cells formed colonies. A similar colony-forming activity was observed in the presence of 10% platelet-poor plasma, but not in the presence of 10% plasma isolated from hypophysectomized rats. Inhibitors of IGF signals attenuated colony formation in the presence of serum. These data suggest that progression factors, probably IGFs, are essential components in serum for ras-induced transformation of 3T3 fibroblasts.     

Complete nucleotide sequence of the human RCC1 gene involved in coupling between DNA replication and mitosis.

Total genomic DNA of the human RCC1 gene was isolated from HeLa DNA and its complete nucleotide sequence (34,641 bp) was determined by the shotgun sequencing method. The exon-intron junctions were precisely assigned to this sequence by comparing the nucleotide sequence of RCC1 genomic DNA with that of its cDNA. The RCC1 gene was found to have 14 exons, 8 of which (starting from the seventh one) coded the seven repeated sequences of RCC1 protein. A single exon corresponded roughly to each repeat of the RCC1 protein except for the middle one, indicating that the RCC1 gene was generated through amplification of a primordial exon. Primer extension analysis revealed the presence of an internal promoter.     

Prevention and reversible solubilization of advanced glycation and products (AGE) by organic germanium compounds as derivatives of amino acids

Summary The amino-carbonyl reaction (The Maillard reaction) of bovine lens crystallin, serum albumin or skin collagen with glucose was investigated to find effective means to prevent the formation of Advanced Glycation End Products (AGE) and induce the reversible solubilization of polymerized glycated proteins. The organic germanium compounds (Ge-132, 373, 385), derivatives of amino acids containing germanium as the linker of framework, were combined by the box titration method to determine the dose that would be most effective, compared with Aminoguanidine-HCl (AMG),-tocopherol (VE), and pirenoxine (Catalin-K, CK). Although AMG suppressed the formation of AGE, effective concentrations were higher than 20 mM. Ge-385, when administered by itself at a low dose, induced the reversible solubilization of AGE made from crystallin, and albumin. The addition of any two reagents such as AMG, VE, CK and Ge-132 or 385 together to proteins lessened the effective range, and the peaks of smaller molecules in the profiles of HPLC and PAGE were quite remarkable. Examination was made of the effects of Ge-132 on the eyes of SAM mice, which show senescence accelerated cataracts at a relatively young age. The prevention of cataract-genesis and induction of reversible transparency of turbid lenses became evident following the administration of Ge-132 to the eyes 4 times a day. The mode of action of organic germanium compounds was demonstrated quite capable of disconnecting the sugar-parts from AGE by decarbonylation, resulting in the formation of glucosone and amino residues, and further leading subsequently to fewer AGE.Abbreviations used in this paper: BLC bovine lens crystallin; BSA bovine serum albumin; AsCol acid soluble bovine skin collagen type III; AGE advanced glycation end products; Ge-132 2-Carboxyethylgermanium sesquioxide, Ge-373 2-Carboxy-2-amino-6-phenyl germanium sesquioxide; Ge-385 2-Carboxy-ethyl-2-aminogermanium sesquioxide; AMG or AG aminoguanidine-HCl; V. E. vitamin E or-tocopherol; CK 1-Hydroxy-5-oxo-5H-pyrido [3, 2-a] phenoxazine-3-carboxylic acid or catalin-K or pirenoxine; PACE polyacrylamide gel electrophoresis; SAM senescence accelerated mouse; HPLC high pressure liquid chromatography; SDS sodium laurylsulfate; FT fructose-p-toluidine.     

Inhibition of Brain Type A Monoamine Oxidase and 5-Hydroxytryptamine Uptake by Two Amphetamine Metabolites, p-Hydroxyamphetamine and p-Hydroxynorephedrine

Two amphetamine metabolites, p-hydroxyamphetamine (p-OHA) and p-hydroxynorephedrine (p-OHN), selectively inhibited the A form of monoamine oxidase (MAO) in rat and mouse forebrain homogenates. Of these two metabolites, p-OHA inhibited MAO-A more strongly than p-OHN. This MAO-A-selective inhibition by p-OHA or p-OHN was found to be competitive with respect to deamination of its substrate, 5-hydroxytryptamine (5-HT). The degree of MAO-A inhibition was not changed by 90 min of preincubation of the enzyme preparations with either metabolite, and the activity inhibited by p-OHA after the preincubation recovered completely to the control level after repeated washing. Uptake of 5-HT or dopamine into mouse forebrain synaptosomes was highly reduced by both p-OHA and p-OHN. Both metabolites were more potent in reducing dopamine uptake than in reducing 5-HT uptake. In reduction of 5-HT and of dopamine uptake, p-OHA was more potent than p-OHN. These results indicate that p-OHA is a more selective inhibitor of brain MAO-A activity and 5-HT uptake than its subsequent metabolite, p-OHN. These two actions of p-OHA might, together with possible 5-HT efflux into the synaptic cleft, greatly contribute to head twitch, a brain 5-HT-mediated animal behavior induced by p-OHA.     

Chromatographic Purification and Determination of the Carboxy-Terminal Sequences of Photosystem II Reaction Center Proteins, Dl and D2

The Dl and D2 subunits of the reaction center of photosystemII are intrinsic proteins, each with a molecular mass of about30 kDa. They exhibit considerable homology to each other interms of primary structure. A procedure was developed for theseparation and purification of these two proteins on a largescale from the photosystem II reaction center complex of spinachby high-performance liquid chromatography on a gel-permeationcolumn in the presence of sodium dodecyl sulfate. The purificationwas achieved by a combination of two gel-permeation chromatographicsteps performed with different concentrations of phosphate buffer,200 mM and 50 mM, as the mobile phase. The purified Dl and D2proteins were subjected to determination of their carboxy-terminalsequences by digestion of the proteins with carboxypeptidaseY. Comparison of the sequences deduced from the enzymatic analysiswith the sequences deduced from the psb A and psb D genes ofspinach indicates that the Dl protein ends at Ala-344 and theD2 protein at Leu-353. Thus, it appears that the Dl proteinloses 9 amino acid residues from the carboxy-terminus, fromAla-345 to Gly-353, during maturation, while the D2 proteindoes not lose any amino acid residues from the carboxy-terminus. (Received July 27, 1989; Accepted December 28, 1989)