BACKGROUND & AIMS: :Upon modification of the reconstituted aspartate/glutamate carrier by mercury reagents the antiporter was converted into a unidirectional efflux carrier (Dierks, T., Salentin, A., Heberger, C. and Krämer, R. (1990) Biochim. Biophys. Acta 1028, 268). In addition to this basic change in the mechanism, the mercurials, reacting with exofacial cysteines, also affected the internal binding site of the carrier leading to an unmeasurable high Km and to a drastically reduced substrate specificity. The spectrum of efflux substrates comprised small anions from chloride to glutamate, but not cationic amino acids and ATP, hence resembling pore-like properties. However, in the efflux state important carrier properties were also observed. The activation energy (86 kJ/mol) was as high as for the antiport. Furthermore, efflux was inhibited by the presence of external substrate. This trans-inhibition strongly suggests that the external binding site of the carrier, prerequisite in the antiport mechanism, also is involved in conformational transitions during efflux function. However, antiport no longer is catalyzed after switching to the efflux state. Reversion of the induced efflux carrier to the antiport state was achieved using dithioerythritol, thereby further restoring substrate specificity and saturation kinetics. A model for antiport-efflux interconversion is presented suggesting that two reactive cysteines have to be modified in order to uncouple the inward and outward directed component of antiport. The pore-type characteristics of efflux are taken as evidence that a channel-like structure determines the selectivity of unidirectional transport. This intrinsic channel of the protein then is required for substrate translocation also during antiport function.

背景与目标： ：在通过汞试剂修饰重构的天冬氨酸/谷氨酸载体后，反转运蛋白转化为单向外排载体（Dierks，T.，Salentin，A.，Heberger，C. andKrämer，R.（1990）Biochim。Biophys。Acta 1028，268）。除了机理上的这一基本变化外，与远颌半胱氨酸反应的汞也影响了载体的内部结合位点，导致无法测量的高Km并大大降低了底物特异性。外排底物的光谱包含从氯离子到谷氨酸的小阴离子，但不包含阳离子氨基酸和ATP，因此具有类似孔的特性。但是，在外排状态下，也观察到重要的载流子性质。活化能（86 kJ / mol）和反端口一样高。此外，外基质的存在抑制了外排。这种反式抑制作用强烈暗示了载体的外部结合位点是反端口机制的先决条件，在外排功能过程中也参与构象转变。但是，切换到外排状态后不再催化反端口。使用二硫赤藓糖醇将诱导的外排载体还原为反端口状态，从而进一步恢复底物特异性和饱和动力学。提出了反端口-外向互转换的模型，该模型建议必须对两个反应性半胱氨酸进行修饰，以解开反端口的向内和向外定向成分。外排的孔型特征被认为是通道状结构决定了单向转运的选择性的证据。然后，在反转运功能期间，该蛋白质的该固有通道也是底物转运所必需的。

BACKGROUND & AIMS: BACKGROUND:In animal models, several anesthetics induce widespread increases in neuronal apoptosis in the developing brain with subsequent neurologic deficits. Although the mechanisms are largely unknown, the neurotoxicity may, at least in part, be due to elevated oxidative stress caused by mitochondrial dysfunction. In an investigation of potential therapies that could protect against this type of damage, we studied the effects of molecular hydrogen on anesthetic-induced neurotoxicity in the developing mouse brain. METHODS:Six-day-old C57BL/6 mice were exposed to 3% sevoflurane for 6 h with or without hydrogen (< 1.3%) as part of the carrier gas mixture. Apoptosis was evaluated by immunohistochemical staining for cleaved caspase-3 (n = 8-10/group). Western blot analysis for cleaved poly-(adenosine diphosphate-ribose) polymerase was also performed to examine apoptosis (n = 3-6/group). Oxidative stress was assessed by immunohistochemical staining for 4-hydroxy-2-nonenal (n = 8/group). Long-term memory and social behavior were examined using the fear conditioning test and the sociability test, respectively (n = 18-20/group). RESULTS:Western blot analysis showed that coadministration of 1.3% hydrogen gas significantly (P < 0.001) reduced the level of neuronal apoptosis to approximately 40% compared with sevoflurane exposure alone. Immunohistochemical analysis showed that hydrogen reduced oxidative stress induced by neonatal sevoflurane exposure. Although neonatal sevoflurane exposure caused impairment in long-term memory and abnormal social behaviors in adulthood, mice coadministered hydrogen gas with sevoflurane did not exhibit these deficits. CONCLUSIONS:Inhalation of hydrogen gas robustly decreased neuronal apoptosis and subsequent cognitive impairments caused by neonatal exposure to sevoflurane.

背景与目标： 背景：在动物模型中，几种麻醉剂会引起发育中的大脑神经元凋亡的广泛增加，并随之导致神经功能缺损。尽管机理尚不清楚，但神经毒性可能至少部分是由于线粒体功能障碍引起的氧化应激升高。在一项可以预防此类损害的潜在疗法的研究中，我们研究了分子氢对麻醉小鼠大脑中麻醉药诱导的神经毒性的影响。
方法：将六日龄的C57BL / 6小鼠暴露于3％的七氟醚中6小时，其中有或没有氢气（<1.3％）作为载气混合物的一部分。通过免疫组织化学染色评估裂解的caspase-3的凋亡（n = 8-10 /组）。还进行了裂解的聚（腺苷二磷酸核糖核糖）聚合酶的蛋白质印迹分析，以检查细胞凋亡（n = 3-6 /组）。通过免疫组织化学染色评估4-羟基-2-壬烯醛的氧化应激（n = 8 /组）。长期记忆和社交行为分别通过恐惧条件测试和社交能力测试（n = 18-20 /组）进行检查。
结果：Western blot分析显示，与单独使用七氟醚相比，共同使用1.3％氢气（P <0.001）可将神经元凋亡水平降低至约40％。免疫组织化学分析表明，氢减少了新生儿七氟醚暴露引起的氧化应激。尽管新生儿七氟醚暴露导致成年期长期记忆受损和社交行为异常，但将氢气与七氟醚并用的小鼠并未表现出这些缺陷。
结论：吸入氢气可有效减少新生儿暴露于七氟醚引起的神经元凋亡和随后的认知障碍。

BACKGROUND & AIMS: PURPOSE:Unverricht-Lundborg disease (EPM1A) is frequently due to an unstable expansion of a dodecamer repeat in the CSTB gene, whereas other types of mutations are rare. EPM1A due to homozygous expansion has a rather stereotyped presentation with prominent action myoclonus. We describe eight patients with five different compound heterozygous CSTB point or indel mutations in order to highlight their particular phenotypical presentations and evaluate their genotype-phenotype relationships. METHODS:We screened CSTB mutations by means of Southern blotting and the sequencing of the genomic DNA of each proband. CSTB messenger RNA (mRNA) aberrations were characterized by sequencing the complementary DNA (cDNA) of lymphoblastoid cells, and assessing the protein concentrations in the lymphoblasts. The patient evaluations included the use of a simplified myoclonus severity rating scale, multiple neurophysiologic tests, and electroencephalography (EEG)-polygraphic recordings. To highlight the particular clinical features and disease time-course in compound heterozygous patients, we compared some of their characteristics with those observed in a series of 40 patients carrying the common homozygous expansion mutation observed at the C. Besta Foundation, Milan, Italy. KEY FINDINGS:The eight compound heterozygous patients belong to six EPM1A families (out of 52; 11.5%) diagnosed at the Laboratory of Genetics of the Galliera Hospitals in Genoa, Italy. They segregated five different heterozygous point or indel mutations in association with the common dodecamer expansion. Four patients from three families had previously reported CSTB mutations (c.67-1G>C and c.168+1_18del); one had a novel nonsense mutation at the first exon (c.133C>T) leading to a premature stop codon predicting a short peptide; the other three patients from two families had a complex novel indel mutation involving the donor splice site of intron 2 (c.168+2_169+21delinsAA) and leading to an aberrant transcript with a partially retained intron. The protein dose (cystatin B/β-actin) in our heterozygous patients was 0.24 ± 0.02, which is not different from that assessed in patients bearing the homozygous dodecamer expansion. The compound heterozygous patients had a significantly earlier disease onset (7.4 ± 1.7 years) than the homozygous patients, and their disease presentations included frequent myoclonic seizures and absences, often occurring in clusters throughout the course of the disease. The seizures were resistant to the pharmacologic treatments that usually lead to complete seizure control in homozygous patients. EEG-polygraphy allowed repeated seizures to be recorded. Action myoclonus progressively worsened and all of the heterozygous patients older than 30 years were in wheelchairs. Most of the patients showed moderate to severe cognitive impairment, and six had psychiatric symptoms. SIGNIFICANCE:EPM1A due to compound heterozygous CSTB mutations presents with variable but often markedly severe and particular phenotypes. Most of our patients presented with the electroclinical features of severe epilepsy, which is unexpected in homozygous patients, and showed frequent seizures resistant to pharmacologic treatment. The presence of variable phenotypes (even in siblings) suggests interactions with other genetic factors influencing the final disease presentation.

背景与目标： 目的：Unrichricht-Lundborg病（EPM1A）通常是由于CSTB基因中十二聚体重复序列的不稳定扩增引起的，而其他类型的突变则很少见。由于纯合子扩展，EPM1A具有刻板印象，表现出明显的肌阵挛作用。我们描述了具有五个不同的复合杂合CSTB点或插入/缺失突变的八位患者，以突出其特定的表型表现并评估其基因型与表型的关系。
方法：我们通过Southern印迹和每个先证者的基因组DNA测序来筛选CSTB突变。 CSTB信使RNA（mRNA）畸变的特征在于，对淋巴母细胞的互补DNA（cDNA）进行测序，并评估淋巴母细胞中的蛋白质浓度。患者评估包括使用简化的肌阵挛严重程度评定量表，多次神经生理学检查和脑电图（EEG）复查记录。为了突出复合杂合患者的特殊临床特征和疾病时程，我们将它们的某些特征与在意大利米兰C. Besta基金会观察到的40例携带普通纯合扩增突变的患者进行了比较。
主要发现：八名复合杂合患者属于六个EPM1A家族（在52个中； 11.5％）是在意大利热那亚Galliera医院的遗传学实验室诊断出的。他们分离了五个不同的杂合点或插入缺失突变，与常见的十二聚体扩展相关。来自三个家庭的四名患者先前曾报告过CSTB突变（c.67-1G> C和c.168 1_18del）；一个在第一个外显子上出现新的无意义突变（c.133C> T），导致提前终止密码子预测短肽。来自两个家庭的其他三名患者具有复杂的新indel突变，涉及内含子2的供体剪接位点（c.168 2_169 21delinsAA），并导致异常转录本和部分保留的内含子。在我们的杂合患者中，蛋白质剂量（胱抑素B /β-肌动蛋白）为0.24±0.02，与纯合十二聚体扩张患者的评估剂量没有差异。复合杂合子患者比纯合子患者显着更早发病（7.4±1.7年），他们的疾病表现包括频繁的肌阵挛性发作和失神，通常在整个病程中成簇出现。癫痫发作对药物治疗有抵抗力，这种治疗通常可导致纯合子患者完全控制癫痫发作。脑电图复查允许记录反复发作。行动性肌阵挛逐渐恶化，所有30岁以上的杂合患者都坐在轮椅上。大多数患者表现出中度至重度认知障碍，其中六人有精神症状。
意义：由于化合物杂合的CSTB突变，EPM1A呈现出可变的但通常具有明显的严重和特殊的表型。我们的大多数患者均表现出严重癫痫的电临床特征，这在纯合子患者中是意料之外的，并且表现出对药物治疗有抗药性的频繁发作。可变表型的存在（甚至在兄弟姐妹中）表明与影响最终疾病表现的其他遗传因素的相互作用。

BACKGROUND & AIMS: :We employed random mutagenesis to determine the region of the initial unfolding of hyper-alkaline-sensitive subtilisin, ALP I, that precedes the denaturation of the entire protein under highly alkaline conditions. This region comprises two alpha-helices and a calcium-binding loop. Stabilization of the region caused the stabilization of the entire protein at a high alkaline pH 12. The alkaline stability of this region was most effectively improved by hydrophobic interactions, followed by ionic interactions with Arg residues. The effect of mutations on the improvement was different with regard to the alkaline stability and thermostability. This indicated that different strategies were necessary to improve the alkaline stability and thermostability of the protein.

背景与目标： ：我们采用随机诱变方法来确定高度碱性敏感的枯草杆菌蛋白酶ALP I的初始展开区域，该区域先于整个蛋白质在高度碱性条件下变性。该区域包括两个α-螺旋和一个钙结合环。该区域的稳定化使整个蛋白质在高碱性pH 12下稳定下来。该区域的碱性稳定性通过疏水性相互作用以及随后与Arg残基的离子相互作用得到了最有效的改善。在碱性稳定性和热稳定性方面，突变对改善的影响是不同的。这表明必须采取不同的策略来提高蛋白质的碱性稳定性和热稳定性。

BACKGROUND & AIMS: :Escherichia coli acyl carrier protein (ACP) contains a single tyrosine residue at position 71. The combined o-nitration of apo-ACP Y71 by tetranitromethane and reduction to 3-aminotyrosyl-apo-ACP were performed to introduce a specific site for attachment of a dansyl fluorescent label. Conditions for purification and characterization of dansylaminotyrosyl-apo-ACP are reported. Dansylaminotyrosyl-apo-ACP was enzymatically phosphopantetheinylated and acylated in vitro with an overall approximately 30% yield of purified stearoyl-dansylaminotyrosyl-ACP starting from unmodified apo-ACP. The steady-state kinetic parameters k(cat) = 22 min(-1) and K(M) = 2.7 microM were determined for reaction of stearoyl-dansylaminotyrosyl-ACP with stearoyl-ACP Delta(9)-desaturase. These results show that dansylaminotyrosyl-ACP will function well for studying binding interactions with the Delta(9)-desaturase and suggest similar possibilities for other ACP-dependent enzymes. The efficient in vivo phosphopantetheinylation of E. coli apo-ACP by coexpression with holo-ACP synthase in E. coli BL21(DE3) using fructose as the carbon source is also reported.

背景与目标： ：大肠杆菌酰基载体蛋白（ACP）在71位含有一个酪氨酸残基。通过四硝基甲烷对apo-ACP Y71进行邻位硝化并还原为3-氨基酪氨酰-apo-ACP，以引入一个特定的位点连接丹磺酰荧光标记。报道了纯化和表征丹磺胺基酪氨酰-apo-ACP的条件。体外将丹磺酰基氨基酪氨酰-Apo-ACP酶促磷酸泛酰化和酰化，从未修饰的载脂蛋白-ACP开始，纯化的硬脂酰基-丹磺酰基氨基酪氨酸-ACP的总收率约为30％。稳态动力学参数k（cat）= 22 min（-1）和K（M）= 2.7 microM被确定为硬脂酰基-丹磺酰基氨基酪氨酰基-ACP与硬脂酰基-ACP Delta（9）-去饱和酶的反应。这些结果表明，dansylaminotyrosyl-ACP将很好地用于研究与Delta（9）-desaturase的结合相互作用，并为其他ACP依赖性酶提出了类似的可能性。还报道了通过使用果糖作为碳源在大肠杆菌BL21（DE3）中与完整ACP合酶共表达来有效表达大肠杆菌apo-ACP的磷酸泛素化。

BACKGROUND & AIMS: :The purpose of this study was to gain a greater understanding of the electrically evoked compound action potential (EAP) responses to pulse train stimulation. Analysis of EAP amplitude responses suggested that an alternating pattern varied depending upon stimulus level, interpulse interval (IPI), stimulus waveform, and stimulus polarity. Stimulus level-dependent recovery was seen in the cat and the guinea pig: higher stimulus level tended to provide faster recovery. Both polarity-dependent recovery and polarity-dependent adaptation were observed in the cat and these stimulus polarity effects were less consistent in the guinea pig. The polarity-dependent recovery effect supports the hypothesis that anodal and cathodal stimuli excite different sites along auditory nerve fibers. Amplitude differences between the response to the second pulse and the steady-state response at the same IPI are significantly greater for anodal stimuli than for cathodal stimuli in all cats. These data suggest that there is a cumulative refractory effect in the auditory nerve of cats, especially in response to anodal stimuli.

背景与目标： ：这项研究的目的是为了更好地了解脉冲诱发的电诱发的复合动作电位（EAP）反应。 EAP振幅响应的分析表明，交替模式会根据刺激水平，脉冲间间隔（IPI），刺激波形和刺激极性而变化。在猫和豚鼠中观察到刺激水平依赖的恢复：较高的刺激水平倾向于提供更快的恢复。在猫中观察到极性依赖性恢复和极性依赖性适应，并且这些刺激性极性效应在豚鼠中不太一致。极性依赖性恢复作用支持以下假设：阳极和阴极刺激会沿听觉神经纤维激发不同的部位。在所有猫中，对于阳极刺激，对第二个脉冲的响应与在相同IPI下的稳态响应之间的幅度差异明显大于对阴极刺激的差异。这些数据表明，猫的听神经具有累积的难治性作用，尤其是对阳极刺激的反应。

BACKGROUND & AIMS: :The bile salt export pump (BSEP) is expressed at the canalicular domain of hepatocytes, where it serves as the primary route of elimination for monovalent bile acids (BAs) into the bile canaliculi. The most compelling evidence linking dysfunction in BA transport with liver injury in humans is found with carriers of mutations that render BSEP nonfunctional. Based on mounting evidence, there appears to be a strong association between drug-induced BSEP interference and liver injury in humans; however, causality has not been established. For this reason, drug-induced BSEP interference is best considered a susceptibility factor for liver injury as other host- or drug-related properties may contribute to the development of hepatotoxicity. To better understand the association between BSEP interference and liver injury in humans, over 600 marketed or withdrawn drugs were evaluated in BSEP expressing membrane vesicles. The example of a compound that failed during phase 1 human trials is also described, AMG 009. AMG 009 showed evidence of liver injury in humans that was not predicted by preclinical safety studies, and BSEP inhibition was implicated. For 109 of the drugs with some effect on in vitro BSEP function, clinical use, associations with hepatotoxicity, pharmacokinetic data, and other information were annotated. A steady state concentration (C(ss)) for each of these annotated drugs was estimated, and a ratio between this value and measured IC₅₀ potency values were calculated in an attempt to relate exposure to in vitro potencies. When factoring for exposure, 95% of the annotated compounds with a C(ss)/BSEP IC₅₀ ratio ≥ 0.1 were associated with some form of liver injury. We then investigated the relationship between clinical evidence of liver injury and effects to multidrug resistance-associated proteins (MRPs) believed to play a role in BA homeostasis. The effect of 600+ drugs on MRP2, MRP3, and MRP4 function was also evaluated in membrane vesicle assays. Drugs with a C(ss)/BSEP IC₅₀ ratio ≥ 0.1 and a C(ss)/MRP IC₅₀ ratio ≥ 0.1 had almost a 100% correlation with some evidence of liver injury in humans. These data suggest that integration of exposure data, and knowledge of an effect to not only BSEP but also one or more of the MRPs, is a useful tool for informing the potential for liver injury due to altered BA transport.

背景与目标： 胆汁盐输出泵（BSEP）在肝细胞的小管结构域表达，是消除单价胆汁酸（BAs）进入胆管的主要途径。发现最令人信服的证据是，BA转运功能障碍与人类肝损伤有关，且携带的突变携带者使BSEP无法起作用。根据越来越多的证据，药物诱发的BSEP干扰与人类肝损伤之间似乎存在很强的联系。但是，因果关系尚未建立。因此，最好将药物诱导的BSEP干扰视为肝损伤的易感因素，因为其他与宿主或药物相关的特性可能会导致肝毒性的发展。为了更好地理解BSEP干扰与人类肝损伤之间的关系，在表达BSEP的膜囊泡中评估了600多种市售或撤回的药物。还描述了在1期人体试验中失败的化合物的示例，即AMG009。AMG009显示了临床前安全性研究未预测到的人类肝损伤的证据，并暗示了BSEP抑制作用。对于109种对体外BSEP功能有一定影响的药物，注明了临床用途，与肝毒性的关系，药代动力学数据和其他信息。估计每种带注释药物的稳态浓度（C（ss）），并计算该值与测得的IC 50效能值之间的比率，以尝试将暴露与体外效能联系起来。当考虑暴露时，C（ss）/ BSEPIC₅₀比≥0.1的95％的带注释化合物与某种形式的肝损伤有关。然后，我们调查了肝损伤的临床证据与对多药耐药相关蛋白（MRP）的影响之间的关系，这些蛋白在BA动态平衡中起作用。还通过膜囊泡试验评估了600种药物对MRP2，MRP3和MRP4功能的影响。 C（ss）/ BSEPIC₅₀比≥0.1和C（ss / MRPIC₅₀比≥0.1）与人类肝损伤的某些证据几乎具有100％的相关性。这些数据表明，暴露数据的整合以及对BSEP以及一个或多个MRP的影响的了解，是一种有用的工具，可用于告知由于BA运输改变而引起肝损伤的可能性。

BACKGROUND & AIMS: :The tubulovesicles of gastric parietal cells sequester H+/K+-ATPase molecules within resting parietal cells. Stimulation of parietal cell secretion elicits delivery of intracellular H+/K+-ATPase to the apically oriented secretory canaliculus. Previous investigations have suggested that this process requires the regulated fusion of intracellular tubulovesicles with the canalicular target membrane. We have sought to investigate the presence of critical putative regulators of vesicle fusion on immunoisolated gastric parietal cell tubulovesicles. Highly purified tubulovesicles were prepared by gradient fractionation and immunoisolation on magnetic beads coated with monoclonal antibodies against the alpha subunit of H+/K+-ATPase. Western blot analysis revealed the presence of Rab11, Rab25, vesicle-associated membrane protein 2 (VAMP-2) and secretory carrier membrane proteins (SCAMPs) on immunoisolated vesicles. The same cohort of proteins was recovered on vesicles immunoisolated with monoclonal antibodies against SCAMPs and VAMP-2. In contrast, whereas immunoreactivities for syntaxin 1A/1B and synaptosome-associated protein (SNAP-25) were present in gradient-isolated vesicles, none of the immunoreactivity was associated with immunoisolated vesicles. The observation of VAMP-2 and two Rab proteins on immunoisolated H+/K+-ATPase-containing tubulovesicles supports the role for tubulovesicles in a regulated vesicle fusion process. In addition, the presence of SCAMPs along with Rab11 and Rab25 implicates the tubulovesicles as a critical apical recycling vesicle population.

背景与目标： ：胃壁细胞的微管壁螯合静息壁细胞内的H / K -ATPase分子。刺激壁细胞分泌引起将细胞内H / K -ATP酶递送至根尖定向的分泌小管。先前的研究表明，该过程需要细胞内微管小泡与小管靶膜的受控融合。我们试图研究在免疫隔离的胃壁细胞微管小泡上囊泡融合的关键推定调节剂的存在。通过梯度分级分离和在涂有针对H / K -ATPase的α亚基的单克隆抗体的磁珠上进行免疫分离来制备高度纯化的微管小泡。蛋白质印迹分析显示免疫隔离的囊泡上存在Rab11，Rab25，囊泡相关膜蛋白2（VAMP-2）和分泌性载体膜蛋白（SCAMPs）。在针对SCAMPs和VAMP-2的单克隆抗体免疫分离的囊泡上回收了同一批蛋白质。相反，尽管在梯度分离的囊泡中存在语法1A / 1B和突触体相关蛋白（SNAP-25）的免疫反应性，但免疫反应性均与免疫分离的囊泡无关。在含有免疫隔离的H / K -ATPase的微管小泡上观察到VAMP-2和两种Rab蛋白支持了微管小泡在调节的囊泡融合过程中的作用。此外，SCAMPs与Rab11和Rab25的存在暗示着微管小泡是一个重要的根尖再生小泡种群。

BACKGROUND & AIMS: :Johanson-Blizzard Syndrome (JBS) is a rare autosomal recessive genetic disorder characterized by exocrine pancreatic insufficiency, distinct abnormal facial appearance and varying degrees of growth retardation. Variants in UBR1 gene are considered to be responsible for the syndrome. Here, we describe a 3-year old boy, who visited our clinic for severe growth retardation and frequent oily diarrhea. The physical examination revealed nasal alae aplasia, scalp defect, and maldescent of left testicle. Transabdominal ultrasound and computed tomography scan of his abdomen demonstrated complete fatty replacement of the pancreas. The clinical, laboratory, and imaging findings strongly suggest the diagnosis of hereditary pancreatitis. Whole exome sequencing revealed two rare compound heterozygous variants, c.2511T > G (p.H837Q) and c.1188T > G (p.Y396X), in the UBR1 gene of this boy, so, the diagnosis of JBS was established. This is the first report of Chinese patient with JBS, and our study indicates that transabdominal ultrasound and computed tomography are two useful and noninvasive imaging methods for the diagnosis and evaluation of JBS, and identification of these two novel variants expands the database of UBR1 gene variants. Furthermore, with the availability of the identification technology for these variants, prenatal diagnosis could be offered for future pregnancies.

背景与目标： ：Johanson-Blizzard综合征（JBS）是一种罕见的常染色体隐性遗传疾病，其特征是外分泌胰腺功能不全，明显的异常面部外观和不同程度的发育迟缓。 UBR1基因的变异被认为是该综合征的原因。在这里，我们描述了一个3岁的男孩，他因严重的发育迟缓和频繁的油性腹泻而去了我们的诊所。体格检查发现鼻阿拉斯发育不全，头皮缺损和左睾丸异常。经腹超声和腹部计算机断层扫描显示胰腺完全脂肪替代。临床，实验室和影像学检查结果强烈提示遗传性胰腺炎的诊断。整个外显子组测序揭示了该男孩的UBR1基因中两个罕见的化合物杂合变异体c.2511T> G（p.H837Q）和c.1188T> G（p.Y396X），因此，JBS的诊断得以确立。这是中国JBS患者的第一份报告，我们的研究表明，经腹超声和计算机断层扫描是诊断和评估JBS的两种有用且无创的成像方法，对这两个新变异的鉴定扩大了UBR1基因变异的数据库。此外，随着这些变体的识别技术的普及，可以为将来的怀孕提供产前诊断。

BACKGROUND & AIMS: :Anophthalmia is a rare eye development anomaly resulting in absent ocular globes or tissue in the orbit since birth. Here, we investigated a newborn with bilateral anophthalmia in a Chinese family. Exome sequencing revealed that compound heterozygous mutations c.287G > A (p.(Arg96His)) and c.709G > A (p.(Gly237Arg)) of the ALDH1A3 gene were present in the affected newborn. Both mutations were absent in all of the searched databases, including 10,000 in-house Chinese exome sequences, and these mutations were confirmed as having been transmitted from the parents. Comparative amino acid sequence analysis across distantly related species revealed that the residues at positions 96 and 234 were evolutionarily highly conserved. In silico analysis predicted these changes to be damaging, and in vitro expression analysis revealed that the mutated alleles were associated with decreased protein production and impaired tetrameric protein formation. This study firstly reported that compound heterozygous mutations of the ALDH1A3 gene can result in anophthalmia in humans, thus highlighting those heterozygous mutations in ALDH1A3 should be considered for molecular screening in anophthalmia, particularly in cases from families without consanguineous relationships.

背景与目标： ：无眼症是一种罕见的眼部发育异常，导致自出生以来眼球或眼眶组织缺失。在这里，我们调查了一个中国家庭的双侧眼球异常的新生儿。外显子组测序显示，在受影响的新生儿中存在ALDH1A3基因的复合杂合突变c.287G> A（p。（Arg96His））和c.709G> A（p。（Gly237Arg））。在所有搜索的数据库中都没有这两个突变，包括10,000个内部中文外显子序列，这些突变被确认是从父母那里传播的。跨远缘物种的比较氨基酸序列分析显示，第96和234位的残基在进化上高度保守。在计算机分析中预测这些变化将是破坏性的，并且体外表达分析显示突变的等位基因与蛋白质产量降低和四聚体蛋白质形成受损有关。这项研究首先报道了ALDH1A3基因的复合杂合突变可导致人类失语，因此强调了ALDH1A3中的那些杂合突变应被用于无眼症的分子筛查，特别是在没有近亲关系的家庭中。

BACKGROUND & AIMS: :Haem carrier protein 1 (HCP1) was originally identified and characterised as a mammalian haem transporter. However, recent evidence has shown that it is also a proton-coupled folate transporter (PCFT) and mutations in the gene cause hereditary folate deficiency in humans. We therefore investigated haem and folate transport characteristics of PCFT/HCP1 both in vivo and in vitro in CD-1 mice and in the presence or absence of a blocking antibody for PCFT/HCP1, and also in cultured cells (which express PCFT/HCP1 endogenously) to elucidate the specificity and selectivity of PCFT/HCP1. The in vivo study showed that the addition of folic acid inhibited 59Fe-labelled haem transport in hypoxic mice but had no effect in normal mice. Using in vitro methods, the results showed increased [3H]folate uptake into everted duodenum from hypoxic mice but uptake was reduced by the addition of haem or PCFT/HCP1 antibodies to the medium. Caco-2 cells transiently transfected with small interfering RNA (siRNA) PCFT/HCP1 duplex oligos resulted in a 69 % reduction in PCFT/HCP1 mRNA when compared with the control siRNA. Both haem and folate uptake were significantly (P < 0.05) reduced in cells transfected with PCFT/HCP1 siRNA; however, the magnitude of reduction with folic acid uptake was greater (48 %) than that of haem (22.5 %). Overall the data support PCFT/HCP1 as a primary folate transporter with a lower affinity for haem. PCFT/HCP1 could therefore play a physiological role in Fe nutrition and the data highlight the potential for the interaction of folate and haem at the level of intestinal absorption.

背景与目标： ：Haem载体蛋白1（HCP1）最初被鉴定为哺乳动物的血红素转运蛋白。但是，最近的证据表明，它也是质子偶联的叶酸转运蛋白（PCFT），该基因的突变会导致人类遗传性叶酸缺乏。因此，我们研究了PC-1 / CD1小鼠体内和体外以及是否存在PCFT / HCP1阻断抗体的情况下，以及培养的细胞（内源性表达PCFT / HCP1）中PCFT / HCP1的血红素和叶酸转运特性）以阐明PCFT / HCP1的特异性和选择性。体内研究表明，添加叶酸可抑制低氧小鼠体内59Fe标记的血红素转运，但对正常小鼠则无作用。使用体外方法，结果表明缺氧小鼠进入外翻十二指肠的[3H]叶酸摄入增加，但通过向培养基中添加血红素或PCFT / HCP1抗体，摄入减少。与对照siRNA相比，用小干扰RNA（siRNA）PCFT / HCP1双链体寡核苷酸瞬时转染的Caco-2细胞导致PCFT / HCP1 mRNA降低了69％。在用PCFT / HCP1 siRNA转染的细胞中，血红素和叶酸的摄取均显着降低（P <0.05）。然而，叶酸摄入量的减少幅度（48％）大于血红素的减少幅度（22.5％）。总体而言，数据支持PCFT / HCP1作为主要的叶酸转运蛋白，对血红素的亲和力较低。因此，PCFT / HCP1可以在铁营养中发挥生理作用，并且数据突显了在肠道吸收水平上叶酸和血红素相互作用的潜力。

BACKGROUND & AIMS: :The genetic and evolutionary relationship among 2,4-diacetylphloroglucinol (Phl)-producing pseudomonads that protect plants from soil-borne pathogens were investigated by multilocus sequence typing. A total of 65 pseudomonads consisting of 58 Phl-positive biocontrol strains of worldwide origin and seven Phl-negative representatives of characterized Pseudomonas species were compared using 10 housekeeping genes (i.e. rrs, dsbA, gyrB, rpoD, fdxA, recA, rpoB, fusA, rpsL and rpsG). Multilocus sequence typing differentiated 51 strains among 58 Phl-positive pseudomonads and proved to be as discriminative as enterobacterial repetitive intergenic consensus polymerase chain reaction profiling. As phylogenetic trees inferred from each locus were rather incongruent with one another, we derived the topology from all concatenated loci, which led to the identification of six main groups of Phl-producing Pseudomonas spp. Taxonomically, these groups could correspond to at least six different species. Linkage disequilibrium analysis pointed to a rather clonal structure, even when the analysis was restricted to Phl-producing pseudomonads from a same geographic location or a same phylogenetic group. Intragenic recombination was evidenced for gyrB, rpoD and fdxA, but was shown to be a weaker force than mutation in the origin of intragenetic diversity. This is the first multilocus assessment of the phylogeny and population structure of an ecologically important bacterial group involved in plant disease suppression.

背景与目标： ：通过多基因座序列分型研究了保护植物免受土壤传播病原体侵害的生产2,4-二乙酰基间苯三酚（Phl）的假单胞菌之间的遗传和进化关系。使用10个管家基因（即rrs，dsbA，gyrB，rpoD，fdxA，recA，rpoB，fusA， rpsL和rpsG）。多基因座序列分型在58个Phl阳性假单胞菌中区分了51个菌株，并被证明与肠细菌重复性基因间共有聚合酶链反应谱具有区别。由于从每个位点推断出的系统发育树彼此之间都不太一致，因此我们从所有串联位点获得了拓扑，从而鉴定了生产Phl的假单胞菌6个主要组。从分类学上讲，这些组可能对应于至少六个不同的物种。连锁不平衡分析指出了相当的克隆结构，即使该分析仅限于来自相同地理位置或相同系统发生群的产Phl的假单胞菌。 gyrB，rpoD和fdxA的基因内重组已得到证实，但被证明比基因内遗传多样性起源中的突变要弱。这是对参与植物病害抑制的重要生态细菌群的系统发育和种群结构的首次多基因座评估。

BACKGROUND & AIMS: :To date, investigations of the hydrophobic substrate site of the insect Delta class glutathione transferase are limited in number. In the present study, putative hydrophobic site residues of AdGSTD4-4 have been proposed and characterized. These residues are Gln-112, Thr-174, Phe-212, Arg-214, Tyr-215 and Phe-216. It was found that Gln-112 does not contribute significantly to the catalytic properties of AdGSTD4-4. Arg-214, Tyr-215 and Phe-216 made contributions to catalytic properties and the rate-limiting step. Thr-174 and Phe-212 appeared to be important in enzymatic catalysis by stabilizing the active site beta1-alpha1 loop on which the critical catalytic residue Ser-9 is located. The aromatic Phe-212 pi cloud appears to be important for interactions with its hydrophobic size representing an almost equally important factor. The data suggests that these residues are not directly involved in catalysis but exert their influence through secondary interactions. In addition, active site rearrangements occur to bring different residues into play even for conjugation through the same mechanisms. Therefore, due to the conformational rearrangements topologically equivalent residues observed in crystal structures may not perform equivalent roles in catalysis in different GST classes.

背景与目标： 迄今为止，对昆虫三角洲谷胱甘肽转移酶的疏水性底物位点的研究数量有限。在本研究中，AdGSTD4-4的假定疏水位点残基已被提出和表征。这些残基是Gln-112，Thr-174，Phe-212，Arg-214，Tyr-215和Phe-216。发现Gln-112对AdGSTD4-4的催化性能没有显着贡献。 Arg-214，Tyr-215和Phe-216对催化性能和限速步骤做出了贡献。通过稳定关键催化残基Ser-9所在的活性位点beta1-alpha1环，Thr-174和Phe-212在酶催化中似乎很重要。芳香族Phe-212π云似乎很重要，因为它的疏水尺寸几乎是同等重要的因素。数据表明这些残基不直接参与催化，而是通过二次相互作用发挥其影响。另外，发生活性位点重排以使不同的残基发挥作用，即使通过相同的机制结合也是如此。因此，由于构象重排，在晶体结构中观察到的拓扑等效残基可能在不同的GST类中的催化作用中不发挥等效作用。

BACKGROUND & AIMS: :Speakers of languages with synchronically productive compounding systems, such as English, are likely to encounter new compounds on a daily basis. These can only be useful for communication if speakers are able to rapidly compose their meanings. However, while compositional meanings can be obtained for some novel compounds such as bridgemill, this is far harder for others such as radiosauce; accordingly, processing speed should be affected by the ease of such a compositional process. To rigorously test this hypothesis, we employed a fully implemented computational model based on distributional semantics to quantitatively measure the degree of semantic compositionality of novel compounds. In two large-scale studies, we collected timed sensibility judgements and lexical decisions for hundreds of morphologically structured nonwords in English. Response times were predicted by the constituents' semantic contribution to the compositional process, with slower rejections for more compositional nonwords. We found no indication of a difference in these compositional effects between the tasks, suggesting that speakers automatically engage in a compositional process whenever they encounter morphologically structured stimuli, even when it is not required by the task at hand. Such compositional effects in the processing of novel compounds have important implications for studies that employ such stimuli as filler material or "nonwords," as response times for these items can differ greatly depending on their compositionality.

背景与目标： ：具有同步生产复合系统的语言（例如英语）的讲者每天可能会遇到新的复合词。这些只有在说话者能够快速表达其含义时才对交流有用。但是，尽管可以对某些新型化合物（例如，bridgemill）获得组成意义，但对其他化合物（如放射性酱）则要困难得多。因此，这种合成方法的容易程度应影响处理速度。为了严格检验该假设，我们采用了一种基于分布语义的完全实现的计算模型，以定量测量新型化合物的语义组成程度。在两项大规模研究中，我们收集了数百种英语中形态结构化的非单词的定时敏感性判断和词汇决策。响应时间是通过成分对组成过程的语义贡献来预测的，对更多组成非单词的拒绝速度较慢。我们没有发现任务之间这些构图效果存在差异的迹象，这表明说话者只要遇到形态结构上的刺激，便会自动参与构图过程，即使手头的任务并不需要。对于使用此类刺激物作为填充材料或“无言词”的研究，这种在新化合物加工中的组成效应具有重要意义，因为这些物品的响应时间可能会因其组成而有很大差异。

BACKGROUND & AIMS: BACKGROUND:Pathogens use multiple mechanisms to disrupt cell functioning in their host and allow pathogenesis. These mechanisms involve communication between the pathogen and the host cell through protein-protein interactions. METHODS:Protein-protein interactions chains referred to as signal transduction pathways are the processes by which a chemical or physical signal transmits through a cell as series of molecular events so the pathogen needs to intercept these molecular pathways at few positions to induce pathogenesis such as pathogen viability, infection or hypersensitivity. RESULTS:The pathogen nodes of interception are not necessarily the most immunogenic; so that novel immunogenicity-improvement strategies need to be developed thought a chemical conjugation of the pathogen-carrier nodes to develop an efficient immune response in order to block pathogenesis. On the other hand, if pathogen-carriers are immunogens; toleration ought to be induced by this conjugation avoiding hypersensitivity. Thus, this paper addresses the biological plausibility of plant-phenolics as pathogen-carrier immunogenicity modulator haptens. CONCLUSION:The plant-phenolic compounds have in their structure functional groups such as hydroxyl, carbonyl, carboxyl, ester, or ether, capable of reacting with the amino or carbonyl groups of the amino acids of a pathogen-carrier to form conjugates. Besides, the varied carbon structures these phenolic compounds have; it is possible to alter the pathogen-carrier related factors that determine the immunogenicity: 1) Structural complexity, 2) Molecular size, 3) Structural heterogeneity, 4) Accessibility to antigenic determinants or epitopes, 5) Optical configuration, 6) Physical state, or 7) Molecular rigidity.

背景与目标： 背景：病原体利用多种机制破坏其宿主中的细胞功能并允许发病。这些机制涉及病原体与宿主细胞之间通过蛋白质-蛋白质相互作用进行的通讯。
方法：蛋白-蛋白质相互作用链被称为信号转导途径，是化学或物理信号作为一系列分子事件通过细胞传递的过程，因此病原体需要在几个位置截断这些分子途径以诱导诸如病原体的发病机理生存力，感染或超敏反应。
结果：截获的病原体不一定具有最强的免疫原性。因此，需要发展新的免疫原性改善策略，考虑到病原体-载体结节的化学结合，以开发有效的免疫应答，从而阻断发病机理。另一方面，如果病原体携带者是免疫原；这种共轭应引起耐受，避免超敏反应。因此，本文探讨了植物酚类作为病原体-载体免疫原性调节剂半抗原的生物学可行性。
结论：植物酚类化合物在结构上具有官能团，例如羟基，羰基，羧基，酯或醚，能够与病原体载体氨基酸的氨基或羰基反应形成结合物。此外，这些酚类化合物具有变化的碳结构。可以改变确定免疫原性的病原体-载体相关因素：1）结构复杂性，2）分子大小，3）结构异质性，4）接近抗原决定簇或表位，5）光学构型，6）物理状态，或7）分子刚性。