Behavioral reactions of rats and the contents of neurospecific proteins in their brain after single X-ray irradiation

We studied the behavior of rats in an open-field test and the contents of neurospecific proteins [neural cell adhesion molecule (NCAM) and glial fibrillary acidic protein (GFAP)] in the brain cortex, hippocampus, striatum, midbrain, cerebellum, andpons Varolii 1, 12, 24, 120, and 168 h after a single X-ray irradiation session (dose of 0.25 Gy). Within the postirradiation period, manifestations of the behavioral activity of the animals were mostly suppressed, and the parameters related to the emotional state of the animals were influenced to a greater extent. The dynamics of the NCAM and GFAP contents were complex and dissimilar in the brain structures under study, but it was possible to observe some general regularities. Within early periods of time, 12 h after irradiation, the NCAM content increased in the cortex, hippocampus, and cerebellum. In these structures, it reached approximately 220, 170, and 150%, respectively, as compared with the control, while it dropped to about 40% in thepons Varolii. Changes in the GFAP content reached their maximum 24 h after irradiation; this index dropped to 29, 44, 34, and 67% in the striatum,pons Varolii, midbrain, and cerebellum, respectively, while it increased to 380% in the hippocampus. Later time intervals were characterized by smoother changes in the contents of the above neurospecific proteins. Seven days after irradiation, the NCAM content did not differ from initial values in the striatum and cerebellum and was higher than the control in the neocortex, hippocampus, and midbrain. Within this period, the GFAP level in the cerebellum and midbrain was relatively normalized, but it increased in the hippocampus and decreased in the pons and striatum. Therefore, the greatest postirradiation shifts in the NCAM and GFAP levels were observed in the structures of the limbic system, and this can be correlated with the data on testing the rats in an open field.     

Effect of Vitamin E on the Content and Polypeptide Composition of Glial Fibrillary Acidic Protein from the Rat Brain under Conditions of Aluminum Chloride Intoxication

We studied the effects of isolated and combined chronic (21 days) introductions of aluminum chloride and vitamin E (α-tocopherol) on the polypeptide composition and content of glial fibrillary acidic protein (GFAP) in different brain structures of rats. Injections of AlCl₃ solution (12 mg/kg, i.p., daily) caused the appearance of low-molecular (47 to 38 kdalton) polypeptides and an increase in the content of GFAP in cytoskeletal fractions to 160 to 220%, as compared with the control. Introduction of vitamin E within the same interval provided significant normalization of the GFAP content in the brain of animals injected with AlCl₃ and to a considerable extent prevented the appearance of degraded polypeptides in the GFAP composition. We discuss the prospects of using vitamin E as an antioxidant for the correction of Al³⁺-induced pathological processes in the CNS.Neirofiziologiya/Neurophysiology, Vol. 37, No. 1, pp. 15–20, January–February, 2005.     

UV-B辐射对拟南芥种子萌发和幼苗生长的影响

以哥伦比亚生态型(Columbia-0)拟南芥(Arabidopsis thaliana)为实验材料, 人工模拟UV-B辐照处理拟南芥种子, 统计其发芽势和发芽率, 测定根长、株高、叶绿素、可溶性糖、可溶性蛋白以及丙二醛(MDA)含量, 研究UV-B辐照处理对拟南芥种子萌发和幼苗生长的影响。研究结果表明, 低剂量的UV-B辐照可以促进拟南芥种子萌发和幼苗生长, 并且最佳辐照剂量为1.0 kJ·m^–2(P<0.05), 此时的发芽势、发芽率、根长、株高、叶绿素、可溶性糖及可溶性蛋白的含量均达到最大, 而丙二醛的含量变化则不明显(P>0.05)。当辐照剂量大于1.0 kJ·m^–2时, 促进作用逐渐变小, 并且随着辐照剂量的增加, 表现出了抑制作用。实验结果表明, 适当剂量的UV-B辐照在一定程度上可以促进拟南芥种子萌发和幼苗生长, 而过高剂量的辐照则对其产生抑制或损伤作用。     

Changes in the Content of Glial Fibrillary Acidic Protein in the Frontal Cortex of Rats during Conditioned Active Avoidance Training

We studied the dynamics of expression of glial fibrillary acidic protein (GFAP) in the frontal neocortex of rats during training for conditioned active avoidance reaction (CAAR). The GFAP content in a cytoskeletal fraction of the frontal cortex tissue was quantitatively estimated with the use of solid-phase immunoenzyme analysis, 2 h and 1, 3, 7, 14, 21, and 30 days after the beginning of training of the animals. It was found that the changes in expression of the filamentous form of GFAP in the neocortex are clearly pronounced and possess a three-phase pattern: the level of GFAP significantly increased within the first hours after the beginning of CAAR training, within 7-14 days this index decreased and became smaller than that in the initial state, and then again rose (with a maximum after 21 days). The obtained data indicate that the neurospecific protein under study is involved in the processes of learning and formation of conditioned memory engrams.     

Response of soil chemistry to forest dieback after bark beetle infestation

We evaluated changes in the chemistry of the uppermost soil horizons in an unmanaged spruce forest (National Park Bohemian Forest, Czech Republic) for 3 years after dieback caused by a bark beetle infestation, and compared these changes with a similar undisturbed forest area. The soils below the disturbed forest received 2–6 times more elements via litter fall compared to the unaffected plot. The subsequent decomposition of litter and reduced nutrient uptake by trees resulted in a steep increase in soil concentrations of soluble N (NH₄-N, organic-bound N) and P forms in the disturbed plot. The average concentrations of NH₄-N and soluble reactive P increased from 0.8 to 4.4 mmol kg^?1 and from 0.04 to 0.9 mmol kg^?1, respectively, in the uppermost soil horizon. Decomposition of litter at the disturbed plot elevated soil concentrations of Ca²⁺, Mg²⁺ and K⁺, which replaced Al³⁺ and H⁺ ions from the soil sorption complex. Consequently, soil concentrations of exchangeable base cations increased from 120 to 200 meq kg^?1, while exchangeable Al³⁺ and H⁺ decreased 66 and 50 %, respectively, and soil base saturation increased from 40 to 70 %. The Al³⁺ liberation did not elevate concentrations of ionic Al in the soil solution, because most of the liberated Al³⁺ was rapidly complexed by dissolved organic carbon (DOC) and transformed to DOC–Al complexes. The chemical parameters investigated at the unaffected plot remained stable during the study.     

Caspase cleavage of GFAP produces an assembly-compromised proteolytic fragment that promotes filament aggregation

IF (intermediate filament) proteins can be cleaved by caspases to generate proapoptotic fragments as shown for desmin. These fragments can also cause filament aggregation. The hypothesis is that disease-causing mutations in IF proteins and their subsequent characteristic histopathological aggregates could involve caspases. GFAP (glial fibrillary acidic protein), a closely related IF protein expressed mainly in astrocytes, is also a putative caspase substrate. Mutations in GFAP cause AxD (Alexander disease). The overexpression of wild-type or mutant GFAP promotes cytoplasmic aggregate formation, with caspase activation and GFAP proteolysis. In this study, we report that GFAP is cleaved specifically by caspase 6 at VELD²²⁵ in its L12 linker domain in vitro. Caspase cleavage of GFAP at Asp²²⁵ produces two major cleavage products. While the C-GFAP (C-terminal GFAP) is unable to assemble into filaments, the N-GFAP (N-terminal GFAP) forms filamentous structures that are variable in width and prone to aggregation. The effect of N-GFAP is dominant, thus affecting normal filament assembly in a way that promotes filament aggregation. Transient transfection of N-GFAP into a human astrocytoma cell line induces the formation of cytoplasmic aggregates, which also disrupt the endogenous GFAP networks. In addition, we generated a neo-epitope antibody that recognizes caspase-cleaved but not the intact GFAP. Using this antibody, we demonstrate the presence of the caspase-generated GFAP fragment in transfected cells expressing a disease-causing mutant GFAP and in two mouse models of AxD. These findings suggest that caspase-mediated GFAP proteolysis may be a common event in the context of both the GFAP mutation and excess.     

The effect of radiation therapy combined with natural killer cells against spontaneous murine fibrosarcoma

The effect of radiation therapy combined with lymphoid cells against spontaneous murine fibrosarcoma (FSa-II) was investigated bothin vivo andin vitro. In thein vivo experiment, syngeneic C3H mice were divided into 3 groups. Animals in the first group were injected with 1 x 10⁵ tumor cells into the right hind leg. Animals in the second and third groups were injected with 1 x 10⁵ tumor cells mixed with 1 x 10⁷ normal lymphoid cells (NLC) or effector lymphoid cells (ELC), respectively. ELC were obtained from spleen and lymph nodes of FSa-II-bearing mice and incubatedin vitro for 40 hr to eliminate suppressor T cell function. NLC were obtained from normal mice and incubated in the same way. Irradiation was given using¹³⁷Cs unit 3 days after cell inoculation. 12 out of 14 mice (85.7%) inoculated with tumor cells mixed with NLC did not show any tumor growth at 60 Gy local irradiation. 12 out of 21 mice (57.1 %) inoculated with tumor cells alone and 6 out of 10 (60%) with tumor cells mixed with ELC rejected tumors at the same radiation dose. This synergistic effect with NLC was not observed when NLC was inoculated after irradiation, indicating that lymphoid cells should be in contact with tumor cells before irradiation. In the⁵¹Cr release assay, lymphoid cells obtained from whole body irradiated (WBI) mice showed 17.8% lysis without irradiation and 28.8% lysis at 5 Gy irradiation. Untreated NLC showed almost no cytotoxic effect at the same radiation dose. This synergistic effect disappeared when WBI lymphoid cells were treated with anti asialo GM1 and complement. These results suggested that NK cells might be important in this synergistic effect with irradiation. To obtain a sufficient level of synergistic effect by in vitro combined treatment of mixed tumor cell - NLC culture and irradiation - incubation for more than 12 hrs and 8 hrs appeared to be necessary before and after irradiation, respectively.     

马尾松幼苗生长及生理特性对铝胁迫的响应

以马尾松幼苗为试验材料,采用水培法研究铝胁迫（Al³⁺浓度为0、0.2、0.4、0.8、1.6 mmol·L^-1）对马尾松幼苗生长及其针叶中叶绿素、渗透调节物质（可溶性糖、可溶性蛋白、脯氨酸）、丙二醛（MDA）和超氧化物歧化酶（SOD）、过氧化物酶（POD）等保护酶活性的影响,为揭示马尾松铝毒害生理机制及提高马尾松的耐铝能力提供理论依据。结果显示：当Al³⁺处理浓度为0.2 mmol·L^-1时对马尾松株高和基径生长的影响较小,但对马尾松根系生长有一定的促进作用;Al³⁺处理浓度大于0.2 mmol·L^-1时对马尾松株高、基径和根长的生长均会产生一定的抑制作用,且这种抑制作用随着Al³⁺浓度的增大而增强。马尾松针叶中叶绿素含量和SOD、POD活性均随着Al³⁺处理浓度的增加呈先上升后下降的趋势;Al³⁺处理浓度大于0.2 mmol·L^-1时马尾松针叶中可溶性糖、可溶性蛋白、脯氨酸等渗透调节物质均呈上升趋势,且随着Al³⁺浓度的升高而增大;Al³⁺处理浓度大于0.2 mmol·L^-1时马尾松针叶中MDA含量也呈上升趋势,且随着Al³⁺浓度的增大而升高,说明大于0.2 mmol·L^-1的Al³⁺处理可导致马尾松膜脂产生氧化。研究表明,马尾松幼苗具有一定的耐铝能力,在铝胁迫生境下可通过提高自身SOD和POD等保护酶的合成和主动积累脯氨酸、蛋白质和可溶性糖等渗透物质,产生适应性生理响应以维持自身的生理平衡来降低铝毒害作用。     

Dynamics of the Bacterial Intermediate Filament Crescentin In Vitro and In Vivo

Background

Crescentin, the recently discovered bacterial intermediate filament protein, organizes into an extended filamentous structure that spans the length of the bacterium Caulobacter crescentus and plays a critical role in defining its curvature. The mechanism by which crescentin mediates cell curvature and whether crescentin filamentous structures are dynamic and/or polar are not fully understood.

Methodology/Principal Findings

Using light microscopy, electron microscopy and quantitative rheology, we investigated the mechanics and dynamics of crescentin structures. Live-cell microscopy reveals that crescentin forms structures in vivo that undergo slow remodeling. The exchange of subunits between these structures and a pool of unassembled subunits is slow during the life cycle of the cell however; in vitro assembly and gelation of C. crescentus crescentin structures are rapid. Moreover, crescentin forms filamentous structures that are elastic, solid-like, and, like other intermediate filaments, can recover a significant portion of their network elasticity after shear. The assembly efficiency of crescentin is largely unaffected by monovalent cations (K⁺, Na⁺), but is enhanced by divalent cations (Mg²⁺, Ca²⁺), suggesting that the assembly kinetics and micromechanics of crescentin depend on the valence of the ions present in solution.

Conclusions/Significance

These results indicate that crescentin forms filamentous structures that are elastic, labile, and stiff, and that their low dissociation rate from established structures controls the slow remodeling of crescentin in C. crescentus.     

Effect of protein deficiency in the female rat diet during pregnancy and lactation on the activity of the membrane and soluble forms of digestive enzymes in the offspring small intestine

Protein deficiency in female rats diet during pregnancy and lactation resulted in deceleration of induction of sucrase both forms in the jejunum and ileum; in acceleration of induction of the maltase membrane from in the jejunum; and in suppression of the lactase membrane form in the ileum; in earlier forming of the adult-type distribution of activity of the membrane form of intestinal alkaline phosphatase and in a decrease in activity of the enzyme soluble form. The findings are corroborated by a suppression of activities of the membrane and soluble forms of the small intestine digestive enzymes in 30-day old rat pups fed with a control (adequate) ration starting 21 days after the birth.     

Synonymous Rare Arginine Codons and tRNA Abundance Affect Protein Production and Quality of TEV Protease Variant

It has been identified that a TEV protease (TEVp) variant, TEVp^5M, displays improved solubility. Here, we constructed fifteen TEVp^5M variants with one or more of six rare arginine codons in the coding sequence replaced with abundant E. coli arginine codons. These codon variants expressed in either E. coli BL21 (DE3) or Rossetta (DE3) showed different solubility and activity. Supply of rare tRNAs increased the tendency of certain codon variants to form insoluble aggregates at early induction stage, as determined by the fused S-tag. About 32% increase in soluble protein production of M5 variant with four synonymously mutated arginine codons was identified in Rossetta (DE3) cells using GFP fusion reporter, comparable to that of TEVp^5M. After purification, two other codon variants from both E. coli strains exhibited less activity than TEVp^5M on cleaving the native or modified recognition sequence incorporated between GST and E. coli diaminopropionate ammonialyase by enzyme-coupled assay, whereas purified M5 variant showed activity similar to the TEVp^5M. Supply of rare tRNAs caused the decrease of activity of TEVp^5M and M5 by about 21%. Our results revealed that engineering of highly soluble TEVp variants can be achieved by the combined mutations of amino acid residues and optimization of specific rare codons, whereas simple augment of rare tRNAs abundance resulted in partial loss of activity.     

Defense responses of soybean roots during exposure to cadmium, excess of nitrogen supply and combinations of these stressors

Heavy metal pollution is a serious environmental problem in agricultural soils since the uptake of heavy metals by plants represents an entry point into the food chain and is influenced by the form and amount of nitrogen (N) fertilization. Here we studied the defense responses in soybean roots exposed to ions of cadmium (applied as 50?mg?l^?1 Cd²⁺) when combined with an excessive dose of N in form of NH₄NO₃. Our data indicate that despite of stunted root growth, several stress symptoms typically observed upon cadmium treatment, e.g. peroxidation of lipid membranes or activation of chitinase isoforms, become suppressed at highly excessive N. At the same time, other defense mechanisms such as catalases and proline accumulation were elevated. Most importantly, the interplay of ongoing responses resulted in a decreased uptake of the metal into the root tissue. This report points to the complexity of plant defense responses under conditions of heavy metal pollution combined with intensive fertilization in agriculture.     

Effect of Aluminum on the Production of Siderophore by Rhizobium sp. (Cicer arietinum)

Rhizobium sp. strain BICC 651 in the presence of 100 μM Al³⁺ produced a threefold higher level of siderophore than in the control culture under iron limitation during the stationary phase. Al³⁺ in increasing concentrations resulted in decreased growth, and the effect was alleviated by the addition of iron. Siderophore production decreased gradually in Al³⁺-treated culture as well as in the control with the addition of increasing concentrations of Fe³⁺, and at 50 μM Fe³⁺ the level of siderophore was practically undetectable. The siderophore binds Fe³⁺ and also Al³⁺. The outer membrane protein profiles of the bacteria grown in the presence or absence of Al³⁺ were indistinguishable. Received: 15 November 1999 / Accepted: 21 December 1999     

Fracto‐mechanoluminescence and thermoluminescence properties of UV and γ‐irradiated Ca2Al2SiO7:Ce3+ phosphor

Ce³⁺‐doped calcium aluminosilicate phosphor was prepared by a combustion‐assisted method at an initiating temperature of 600°C. Structural characterization was carried out using X‐ray diffraction (XRD) and scanning electron microscopy (SEM). The absorption spectra of Ca₂Al₂SiO₇:Ce³⁺ showed an absorption edge at 230 nm. The optical characterization of Ca₂Al₂SiO₇:Ce³⁺ phosphor was investigated in a fracto‐mechanoluminescence (FML) and thermoluminescence (TL) study. The peak of ML intensity increased as the height of impact of the moving piston increased. The TL intensity of Ca₂Al₂SiO₇:Ce³⁺ was recorded for different exposure times of UV and γ‐irradiation and it was observed that TL intensity was maximum for a UV irradiation time of 30 min and for a γ‐dose of 1180 Gy. The TL intensity had three peaks for UV irradiation at temperatures 82°C, 125°C and 203°C. Also the TL intensity had a single peak at 152°C for γ‐irradiation. The TL and ML emission spectra of Ca₂Al₂SiO₇:Ce³⁺ phosphor showed maximum emission at 400 nm. The possible mechanisms involved in the TL and ML processes of the Ca₂Al₂SiO₇:Ce³⁺ phosphor are also explained. Copyright © 2015 John Wiley & Sons, Ltd.     

Involvement of oxidative stress and role of antioxidative defense system in growing rice seedlings exposed to toxic concentrations of aluminum

When seedlings of rice (Oryza sativa L.) cultivar Pant-12 were raised in sand cultures containing 80 and 160 μM Al³⁺ in the medium for 5–20 days, a regular increase in Al³⁺ uptake with a concomitant decrease in the length of roots as well as shoots was observed. Al³⁺ treatment of 160 μM resulted in increased generation of superoxide anion (O₂ ⁻) and hydrogen peroxide (H₂O₂), elevated amount of malondialdehyde, soluble protein and oxidized glutathione and decline in the concentrations of thiols (-SH) and ascorbic acid. Among antioxidative enzymes, activities of superoxide dismutase (SOD EC 1.15.1.1), guaiacol peroxidase (Guaiacol POX EC 1.11.1.7), ascorbate peroxidase (APX EC 1.11.1.11), monodehydroascorbate reductase (MDHAR EC 1.6.5.4), dehydroascorbate reductase (EC 1.8.5.1) and glutathione reductase (EC 1.6.4.2) increased significantly, whereas the activities of catalase (EC EC 1.11.1.6) and chloroplastic APX declined in 160 μM Al³⁺ stressed seedlings as compared to control seedlings. The results suggest that Al³⁺ toxicity is associated with induction of oxidative stress in rice plants and among antioxidative enzymes SOD, Guaiacol POX and cytosolic APX appear to serve as important components of an antioxidative defense mechanism under Al³⁺ toxicity. PAGE analysis confirmed the increased activity as well as appearance of new isoenzymes of APX in Al³⁺ stressed seedlings. Immunoblot analysis revealed that changes in the activities of APX are due to changes in the amounts of enzyme protein. Similar findings were obtained when the experiments were repeated using another popular rice cv. Malviya-36.     

Aluminum‐dependent dynamics of ion transport in Arabidopsis: specificity of low pH and aluminum responses

Low‐pH and Al³⁺ stresses are the major causes of poor plant growth in acidic soils. However, there is still a poor understanding of plant responses to low‐pH and Al³⁺ toxicity. Low‐pH or combined low‐pH and Al³⁺ stress was imposed in order to measure rhizosphere pH, ion fluxes, plasma membrane potential and intracellular H⁺ concentration in distal elongation and mature zones (MZs) along the longitudinal axis of Arabidopsis thaliana roots. Low‐pH stress facilitated H⁺ influx into root tissues and caused cytoplasmic acidification; by contrast, combined low‐pH/Al³⁺ treatment either decreased H⁺ influx in the distal elongation zone (DEZ) or induced H⁺ efflux in the MZ, leading to cytoplasmic alkalinization in both zones. Low‐pH stress induced an increase in rhizosphere pH in the DEZ, whereas combined low‐pH/Al³⁺ stress resulted in lower rhizosphere pH in both root zones compared with the low‐pH treatment alone. Low‐pH stress facilitated K⁺ efflux; the presence of Al³⁺ diminished K⁺ efflux or favored K⁺ influx into root tissues. In both zones, low‐pH treatment induced plasma membrane (PM) depolarization, which was significantly diminished (P≤ 0.05) when combined stresses (low‐pH/100 µM Al³⁺) were imposed. After 60 min of exposure, low pH caused PM depolarization, whereas low pH/100 µM Al³⁺ caused PM hyperpolarization. Thus, low pH and Al³⁺ toxicity differentially affect root tissues and, consequently, the rhizosphere, which might underpin the differential mechanisms of plant adaptation to these abiotic stresses.     

Thermoluminescent properties of some phosphor glasses based on aluminum oxide

The transparent thermoluminescent aluminum oxide‐based glass of 15Al₂O₃–35P₂O₅–25CaO–25Na₂CO₃, abbreviated as APCN (all in mol%) doped with different concentrations of SiO₂ from 0.0–500 ppm was prepared using a conventional melt‐quenching technique. The TL sensitivities of the prepared glasses were investigated at 3 Gy γ‐dose using a ⁶⁰Co source and measured at a heating rate 10 C/sec. The highest TL intensity of the material doped with SiO₂ was found at a concentration of 500 ppm (APCNSi₅). Deconvolution of the glow curve from APCNSi₅ resulted in four peaks at about 161, 194, 237 and 293 C with a Figure Of Merit (FOM) of 1.28%. The APCNSi₅ specimen had the best dosimetric properties when compared with the other samples. Reproducibility, repeatability, dose–response curve and fading effect were checked for peak 3, which appeared at about 237 C. The results displayed that the APCNSi₅ glass system was a low‐Z material (Z_eff ≈ 10), and had good reproducibility and good repeatability. Peak 3 showed good linearity over a dose range up to 20 Gy (R² = 0.999) and sublinearity behaviour was found. The signal from APCNSi₅ faded by about 11% after 2 days post irradiation, therefore it showed almost no significant loss. Such properties make the newly prepared glasses suitable for and highly recommended for use in γ‐dosimeters. Copyright © 2016 John Wiley & Sons, Ltd.     

柠檬香蜂草对铜的耐性及其积累特征研究

以柠檬香蜂草(Melissa officinalis)幼苗为材料,设置不同浓度Cu~(2+)胁迫(CK、200、400、800和1 000mg·kg~(-1))盆栽实验,测定胁迫0、7、14、21、28d后植物生物量、叶绿素含量、抗氧化酶活性、可溶性蛋白含量、丙二醛(MDA)含量以及植物体内Cu含量等指标,探讨柠檬香蜂草对Cu~(2+)的耐受性及其积累特征。结果表明:(1)相同处理时间下,柠檬香蜂草除MDA含量外其他所有指标均随着Cu~(2+)胁迫处理浓度的增加呈低浓度促进、高浓度抑制的变化趋势,且高浓度组(800和1 000mg·kg~(-1))与低浓度组(200和400mg·kg~(-1))之间差异显著(P0.05);MDA含量在1 000mg·kg~(-1)浓度下持续增长至第14天后开始下降。(2)柠檬香蜂草体内Cu的积累量随Cu胁迫浓度的升高呈先增加后减少的趋势,并在浓度为400mg·kg~(-1)时达到最高值(0.71mg/盆)。(3)在整个胁迫过程中,柠檬香蜂草植株的铜富集系数及其耐性系数均随Cu浓度的增加而减小,各处理浓度对Cu的耐性系数均大于0.5,富集系数均大于1。研究发现,柠檬香蜂草对Cu胁迫具有一定耐受性和富集能力,具有成为铜污染土壤修复植物的潜力。     

Effect of manganese and other heavy metals on submerged citric acid fermentation of molasses

The addition of as little as 2 ppb of manganese to ferrocyanide-treated beet molasses during citric acid fermentation by Aspergillus niger NRC A-1-233 caused a 10% reduction in acid yield and undesirable change in the morphology of the organism from the normal pelletlike form to the filamentous from. Still smaller additions (0.4-2ppb) caused undesirable pellet clumping, while greater additions (2-100 ppb) gave further decreases in yield. The yield obtained at 100 ppb was less than 25% of that obtained at 1 ppb or less. None of the other metals tested (Al³⁺, Ca²⁺, Co²⁺, Cu²⁺, Fe²⁺, Mg²⁺, Ni²⁺, Zn²⁺) visibly changed pellet morphology, and only Al³⁺, Fe²⁺, and Zn²⁺ at relatively higher concentrations (5–25ppm) reduced acid yield. The adverse effect of manganese on growth and acid production was not affected by addition of the other metals.     

Oxytocin solution structure changes upon protonation of the N-terminus in dimethyl sulfoxide

Summary With the combined use of various two-dimensional (2D) NMR techniques, a complete assignment of the ¹H and ¹³C resonances of oxytocin, , for two molecular states, protonated and unprotonated at the N-terminal group, was performed in dimethyl sulfoxide. A small but distinct change in the backbone conformation of the six-residue cyclic moiety, associated with the protonation, was first suggested from those NMR parameters relevant to conformation, such as change with temperature in the chemical shifts of the peptide amide protons and changes in chemical shifts and homonuclear as well as heteronuclear three-bond coupling constants. The solution structures of oxytocin for the protonated and unprotonated forms were then calculated using distance analysis in dihedral-angle space, based on a relaxation matrix evaluated from quantitative NOE intensities at different mixing times. Total amounts of 93 and 105 distances were determined for the protonated and the unprotonated forms, respectively. There were 25 interresidue distances relevant to the structure of the cyclic moiety for the protonated form of oxytocin and 43 for the unprotonated form. Overall structures with the lowest target penalty function were similar between the two forms, having a -turn structure at the endocyclic residues of the Tyr-Ile-Gln-Asn moiety. The local backbone conformations near the N-terminus, however, were significantly different between the two forms. This was found to be due to a change in the dihedral angle of the disulfide bridge (^ss around C-S-S-C), which closes the ring in the cyclic peptide. The dihedral angle was about +90° for the unprotonated form and an intermediate value of about +45° for the protonated form.