抗菌肽改良设计及抗炎作用的研究进展

抗菌肽因其具有广谱抗菌活性、不容易引起抵抗性,被认为是先天免疫系统对抗微生物感染的多功能工具。然而,天然抗菌肽存在抗菌活性低、稳定性低、溶血性高等问题,使其较难应用于临床,所以研究人员对抗菌肽进行改良设计以期获得更高抗菌活性、更低溶血活性的新型抗菌肽。另外,天然抗菌肽作为一类免疫效应因子而被发现,其表现出的抑菌、免疫调节、内毒素中和等作用,使得研究人员对抗菌肽在抗炎作用的研究表现出极大的兴趣。就抗菌肽的药物设计方法及抗炎作用机制进行综述。     

Aromatic l-Amino Acid Decarboxylase Activity of the Rat Retina Is Modulated In Vivo by Environmental Light Maria Hadjiconstantinou

Aromatic L-amino acid decarboxylase (AAAD) activity of rat retina is low in animals placed in the dark. When the room lights are turned on, activity rises for almost 3 h and reaches values that are about twice the values found in the dark. A study of the kinetics of the enzyme revealed that the apparent Km values for L-3,4-dihydroxyphenylalanine and pyridoxal-5'-phosphate were unchanged in light- and dark-exposed animals, whereas the Vmax increased in the light. Treating the animals with cycloheximide before exposure to light prevented the increase of enzyme activity. Immunotitration with antibodies to AAAD suggested that more enzyme molecules are present in the light than in the dark. When the room lights are turned off AAAD activity drops rapidly at first and then more slowly, suggesting that at least two processes are responsible for the fall of enzyme activity. Exposure to short periods of dark followed by light results in a rapid increase of AAAD activity. Mixing homogenates from light- and dark-exposed rats results in activity values that are less than expected, suggesting the presence of an endogenous inhibitor(s). These studies demonstrate that AAAD activity is modulated in vivo.     

Activity-dependent clustering of functional synaptic inputs on developing hippocampal dendrites

During brain development, before sensory systems become functional, neuronal networks spontaneously generate repetitive bursts of neuronal activity, which are typically synchronized across many neurons. Such activity patterns have been described on the level of networks and cells, but the fine-structure of inputs received by an individual neuron during spontaneous network activity has not been studied. Here, we used calcium imaging to record activity at many synapses of hippocampal pyramidal neurons simultaneously to establish the activity patterns in the majority of synapses of an entire cell. Analysis of the spatiotemporal patterns of synaptic activity revealed a fine-scale connectivity rule: neighboring synapses (<16?μm intersynapse distance) are more likely to be coactive than synapses that are farther away from each other. Blocking spiking activity or NMDA receptor activation revealed that the clustering of synaptic inputs required neuronal activity, demonstrating a role of developmentally expressed spontaneous activity for connecting neurons with subcellular precision.     

Intact plastids are required for nitrate- and light-induced accumulation of nitrate reductase activity and mRNA in squash cotyledons

Induction of nitrate reductase activity and mRNA by nitrate and light is prevented if chloroplasts are destroyed by photooxidation in norflurazon-treated squash (Cucurbita maxima L.) cotyledons. The enzyme activity and mRNA can be induced if norflurazon-treated squash seedlings are kept in low-intensity red light, which minimizes photodamage to the plastids. It is concluded that induction of nitrate reductase activity and nitrate reductase mRNA requires intact plastids. If squash seedlings grown in low-intensity red light are transferred to photooxidative white light, nitrate reductase activity accumulates during the first 12 hours after the shift and declines thereafter. Thus photodamage to the plastids and the disappearance of nitrate reductase activity and mRNA are events separable in time, and disappearance of the enzyme activity is a consequence of the damage to the plastids.     

龟类野外活动节律的研究进展

龟类野外活动具有时间尺度上的差异性和周期性,温度、湿度、光照、天气、年龄和性别差异等各种因子对龟的野外活动有着不同程度的影响.从日活动、季节活动两个不同时间尺度对龟类野外活动节律的研究成果做了系统总结,将休眠作为一种特殊的季节活动进行了小结,并对影响龟类野外活动节律的相关因子进行了分析和探讨.     

Bacillus stearothermophilus PcrA monomer is a single-stranded DNA translocase but not a processive helicase in vitro

Structural studies of the Bacillus stearothermophilus PcrA protein along with biochemical studies of the single-stranded (ss) DNA translocation activity of PcrA monomers have led to the suggestion that a PcrA monomer possesses processive helicase activity in vitro. Yet definitive studies testing whether the PcrA monomer possesses processive helicase activity have not been performed. Here we show, using single turnover kinetic methods, that monomers of PcrA are able to translocate along ssDNA, in the 3' to 5' direction, rapidly and processively, whereas these same monomers display no detectable helicase activity under the same solution conditions in vitro. The PcrA monomer ssDNA translocation activity, although necessary, is not sufficient for processive helicase activity, and thus the translocase and helicase activities of PcrA are separable. These results also suggest that the helicase activity of PcrA needs to be activated either by self-assembly or through interactions with accessory proteins. This same behavior is displayed by both the Escherichia coli Rep and UvrD monomers. Hence, all three of these SF1 enzymes are ssDNA translocases as monomers but do not display processive helicase activity in vitro unless activated. The fact that the translocase and helicase activities are separable suggests that each activity may be used for different functions in vivo.     

Identification of a deoxyribonuclease activity in the nuclei of the cervical carcinoma HeLa

A deoxyribonuclease activity has been identified in the nuclei of the human cervical carcinoma HeLa. This activity has the novel property of existing as a single strand specific endo- and exodeoxyribonuclease activity. These activities are ionic strength sensitive, with retardation of activity occurring at 50 mM NaCl and above, with the Mn++ ion preferred over the Mg++ ion as activating cation. This activity extensively damages DNA via its single strand nicking and gaping activity. The method used to solubilize this activity as well as the enzyme's characteristics are discussed.     

Defying the activity–stability trade-off in enzymes: taking advantage of entropy to enhance activity and thermostability

The biotechnological applications of enzymes are limited due to the activity–stability trade-off, which implies that an increase in activity is accompanied by a concomitant decrease in protein stability. This premise is based on thermally adapted homologous enzymes where cold-adapted enzymes show high intrinsic activity linked to enhanced thermolability. In contrast, thermophilic enzymes show low activity around ambient temperatures. Nevertheless, genetically and chemically modified enzymes are beginning to show that the activity–stability trade-off can be overcome. In this review, the origin of the activity–stability trade-off, the thermodynamic basis for enhanced activity and stability, and various approaches for escaping the activity–stability trade-off are discussed. The role of entropy in enhancing both the activity and the stability of enzymes is highlighted with a special emphasis placed on the involvement of solvent water molecules. This review is concluded with suggestions for further research, which underscores the implications of these findings in the context of productivity curves, the Daniel–Danson equilibrium model, catalytic antibodies, and life on cold planets.     

Bordetella pertussis adenylate cyclase toxin. Structural and functional independence of the catalytic and hemolytic activities.

The Bordetella pertussis calmodulin-dependent adenylate cyclase (CyaA) is a 1706-residue-long toxin, endowed with hemolytic activity. We have constructed B. pertussis mutant strains producing modified CyaAs devoid of adenylate cyclase activity. Our results show that such modified CyaAs display hemolytic activity identical to the wild-type toxin, thus demonstrating that the hemolytic activity is independent of the adenylate cyclase activity. Furthermore, B. pertussis and Escherichia coli strains producing CyaA lacking the catalytic domain (residues 1-373) were constructed. The truncated protein exhibits hemolytic activity comparable to the wild-type toxin, thus establishing that the carboxyl-terminal 1332 residues alone are endowed with hemolytic activity. Together, these findings show that adenylate cyclase and hemolytic activities are located in two distinct regions of the molecule (respectively, approximately amino acids 1-400 and 401-1706) and that the two regions of CyaA are functionally independent.     

河西走廊芦苇的光合碳同化途径对生境条件的适应

 本文以甘肃省河西走廊生长的四个不同生境芦苇为对象,比较研究了它们的叶解剖结构、光合关键酶活力、乙醇酸氧化酶活力和稳定碳同位素组成(δ13C)。结果发现,沼泽芦苇叶中虽具有不典型的花环结构,但维管束鞘细胞中不含叶绿体,RUBPcase活力／PEPcase活力比值为24.4,乙醇酸氧化酶活力为1218Unit mgpro-1·min-1δ13C值为-34‰,这些测值位于C3植物(小麦)的范围内。生长于沙丘上的芦苇叶片具有明显的花环结构,维管束鞘细胞内含异型叶绿体,RUBPcase活力／PEPcase活力比值为0.985, 乙醇酸氧化酶活力为504 Unitmgpro-1·min-1,δ13C值为-20.9‰,这些测值与典型C4植物(玉米)十分相似。盐化草甸芦苇和盐化草一沙丘过渡地带芦苇叶中均具有明显的花环结构,维管束鞘细胞中含大型叶绿体,RUBPcase活力／PEPcase活力比值分别为2.45和1.53,但乙醇酸氧化酶活力分别为1470和2058Unitmgpro-1·min-1,δ13C值分别为35.6和30.6‰,综合盐化草甸芦苇和过渡地带芦苇的上述指标,似介于沼泽芦苇和沙丘芦苇之间。由这些结果可以认为,分布于甘肃省河西走廊的芦苇,在种内发生有由环境因子引起的光合碳代谢途径的适应性改变。     

Ultrastructural distribution of peroxidase in thrombocytes of mammals and submammals

The localization and distribution of peroxidase (PPO) activity were studied ultracytochemically in thrombocytes from lampreys, carps, frogs, snakes, tortoises, rabbits, sheep, dogs, and monkeys. PPO activity was not detectable in the thrombocytes of lampreys, carps, frogs, and snakes. However, this enzyme activity was demonstrated in the nuclear envelope and endoplasmic reticulum of tortoise thrombocytes. Dog and monkey thrombocytes (blood platelets) exhibited PPO activity in the dense tubular system, but this enzyme activity was not detectable in rabbit and sheep thrombocytes. Our observations are interpreted to suggest that thrombocytes from animals lower than amphibia are peroxidase negative. Furthermore, it can be said that thrombocytes from animals higher than reptiles are generally positive, although there are exceptions. PPO activity was localized in the endoplasmic-reticulum system, but not in the cytoplasmic granules of thrombocytes common to submammals and mammals. In this study, we also compared the distribution of peroxidase activity in thrombocytes, neutrophils, and eosinophils and conclude that these are significant differences in the distribution of PPO and myeloperoxidase.     

Distribution of activities of aspartate aminotransferase isoenzymes and malate dehydrogenase in guinea pig retinal layers

Distributions of activity of the cytosolic (cAAT) and mitochondrial (mAAT) isoenzymes of aspartate aminotransferase and of malate dehydrogenase (MDH) were determined in guinea pig retinal layers. The distribution of total AAT activity (tAAT = cAAT + mAAT) and of mAAT activity correlated well (r = 0.88-0.91) with the distribution of MDH activity. mAAT activity was highest in the inner segments of the photoreceptors; there was a greater than twelve-fold difference between activity in that layer and in the inner retinal layers. cAAT activity was also highest in the inner segments, but the difference between the activity in the inner segments and the other layers was not nearly as great as with mAAT. cAAT activity was also relatively high in the outer nuclear layer, outer plexiform layer, and part of the inner plexiform layer. The high activity of cAAT, mAAT, and MDH in the inner segments indicates that all of these enzymes are involved in metabolic reactions related to energy production and/or to photoreceptive processes in the outer segments and, therefore, that the enzymes are probably involved in energy-related metabolism at synapses. However, other functions, including those related to neurotransmission, are not excluded.     

Observing Single Enzyme Molecules Interconvert between Activity States upon Heating

In this paper, we demonstrate that single enzyme molecules of β-galactosidase interconvert between different activity states upon exposure to short pulses of heat. We show that these changes in activity are the result of different enzyme conformations. Hundreds of single β-galactosidase molecules are trapped in femtoliter reaction chambers and the individual enzymes are subjected to short heating pulses. When heating pulses are introduced into the system, the enzyme molecules switch between different activity states. Furthermore, we observe that the changes in activity are random and do not correlate with the enzyme''s original activity. This study demonstrates that different stable conformations play an important role in the static heterogeneity reported previously, resulting in distinct long-lived activity states of enzyme molecules in a population.     

The daily activity rhythm in a troop of wild Japanese monkey

An assessment of the daily activity rhythm of wild Japanese monkeys was tried both from the calculation of the proportion that each activity occupied in the total activities and the “nomadograph,” representing temporary change in the pace of the daily movement. Seasonal and day-to-day changes are recognized in the daily activity rhythm of the troop of wild Japanese monkeys. It seems that seasonal change in the daily activity rhythm corresponds to the seasonal fluctuation of food supply and atmospheric temperature. From autumn to early winter, when much food is available, a clear-cut pattern of activity emerges; namely, three intensive feeding periods are recognized in a day. Moreover, day-to-day variation in the activity rhythm is fairly small and the activity pattern thus becomes standardized. In winter, when least food is available, activity of monkeys drops to the lowest level of the year. Day-to-day variation in the activity rhythm is great. Two to four intensive feeding periods in a day are recognized. In early spring and summer, when food supply is rather scarce, there exist two to three intensive feeding periods in a day. During the heat of the day in summer, activity of monkeys is conspicuously low.     

Receptor Level Mechanisms Are Required for Epidermal Growth Factor (EGF)-stimulated Extracellular Signal-regulated Kinase (ERK) Activity Pulses

In both physiological and cell culture systems, EGF-stimulated ERK activity occurs in discrete pulses within individual cells. Many feedback loops are present in the EGF receptor (EGFR)-ERK network, but the mechanisms driving pulsatile ERK kinetics are unknown. Here, we find that in cells that respond to EGF with frequency-modulated pulsatile ERK activity, stimulation through a heterologous TrkA receptor system results in non-pulsatile, amplitude-modulated activation of ERK. We further dissect the kinetics of pulse activity using a combination of FRET- and translocation-based reporters and find that EGFR activity is required to maintain ERK activity throughout the 10–20-minute lifetime of pulses. Together, these data indicate that feedbacks operating within the core Ras-Raf-MEK-ERK cascade are insufficient to drive discrete pulses of ERK activity and instead implicate mechanisms acting at the level of EGFR.     

Reconstruction of behavior-relevant individual brain activity: an individualized fMRI study

Different patterns of brain activity are observed in various subjects across a wide functional domain.However,these individual differences,which are often neglected through the group average,are not yet completely understood.Based on the fundamental assumption that human behavior is rooted in the underlying brain function,we speculated that the individual differences in brain activity are reflected in the individual differences in behavior.Adopting 98 behavioral measures and assessing the brain activity induced at seven task functional magnetic resonance imaging states,we demonstrated that the individual differences in brain activity can be used to predict behavioral measures of individual subjects with high accuracy using the partial least square regression model.In addition,we revealed that behavior-relevant individual differences in brain activity transferred between different task states and can be used to reconstruct individual brain activity.Reconstructed individual brain activity retained certain individual differences which were lost in the group average and could serve as an individual functional localizer.Therefore,our results suggest that the individual differences in brain activity contain behavior-relevant information and should be included in group averaging.Moreover,reconstructed individual brain activity shows a potential use in precise and personalized medicine.     

Muscle fibre composition of the eye muscles of the carp (Cyprinus carpio L.) defined on the base of histochemical observations

A histochemical study has been carried out on the eye muscles of the carp. On the base of the ATP-ase and SDH activity and with regard to the localization and diameter of muscle fibres six types of muscle fibres situated in the defined zones can be distinguished. The type 1 and 2 fibres display a moderately high ATP-ase activity at pH 9.4 and rather low activity after alkaline and acid preincubation. Type 1 shows a high SDH activity in contrast to type 2 with a low SDH activity. The other four types of fibres have the high ATP-ase activity at pH 9.4. Type 3 contains fibres with a moderately high ATP-ase activity after alkaline preincubation with a rather low activity after acid preincubation and with a low SDH activity. The fibres of type 4 characterized by the high ATP-ase activity after alkaline and acid preincubation and by the high SDH activity. The fibres of type 5 display high ATP-ase activity after alkaline and acid preincubation and the low SDH activity. They are situated in the white and intermediate fibre zones. The fibres of type 6 are comparable to the fibres of type 5, however they differ diameter and localization, i.e. they are situated in small diameter fibre zone. Using the electron microscope four types of fibres (A, B, C, and D) are found. They vary in the localization of T system, the organization of Z-line and in M-line appearance. In the type B of muscle fibres two different localizations of T system have been discerned.     

高原鼠兔似昼夜活动节律的研究

本文对高原鼠兔活动节律的季节变化,不同光强对高原鼠兔似昼夜活动节律的影响,以及温度效应,饥俄效应等问题进行了研究。同时对昼夜节律研究中长期存在的理论问题,内源性和外源性观点提出探讨。     

Polyphasic activity patterns in small mammals

Cathemeral species are routinely active during the day, the night and at twilight. For the majority of species it is advantageous to specialize on the environmental conditions of a particular phase of the 24-hour day, so this rather uncommon type of activity must be a consequence of specific constraints. Good examples are the polyphasic activity patterns found in some small mammals. In shrews, with small body size and extremely high metabolic rate, polyphasic activity represents a simple short-term hunger cycle. In voles the short-term rhythm is triggered by an additional endogenous ultradian clock that interacts with the common circadian system, which probably is functionally related to endosymbiont digestion of cellulose-rich food. The activity bouts of individuals are synchronized on the population level to spread predation risk. As cathemeral species, voles are not specifically adapted to particular light conditions, but they are also not restricted to a particular activity phase. Therefore, the benefits from flexible responses in activity timing to environmental challenges may compensate for the disadvantages of not being specialized.