Selection of trkB-binding peptides from a phage-displayed random peptide library

Brain-derived neurotrophic factor (BDNF) shows potential in the treatment of neurodegenerative diseases, but the therapeutic application of BDNF has been greatly limited because it is too large in molecular size to permeate blood-brain barrier. To develop low-molecular-weight BDNF-like peptides, we selected a phage-displayed random peptide library using trkB expressed on NIH 3T3 cells as target in the study. With the strategy of peptide library incubation with NIH 3T3 cells and competitive elution with 1 μg/mL of BDNF in the last round of selection, the specific phages able to bind to the natural conformation of trkB and antagonize BDNF binding to trkB were enriched effectively. Five trkB-binding peptides were obtained, in which a core sequence of CRA/TXΦXXΦXXC (X represents the random amino acids, Φ represents T, L or I) was identified. The BDNF-like activity of these five peptides displayed on phages was not observed, though all of them antagonized the activity of BDNF in a dose-dependent manner. Similar results were obtained with the synthetic peptide of C1 clone, indicating that the 5 phage-derived peptides were trkB antagonists. These low-molecular-weight antagonists of trkB may be of potential application in the treatment of neuroblastoma and chronic pain. Meanwhile, the obtained core sequence also could be used as the base to construct the secondary phage-displayed peptide library for further development of small peptides mimicking BDNF activity.     

Selection of trkB-binding peptides from a phage-displayed random peptide library

Brain-derivedneurotrophicfactor(BDNF),originallypurifiedfrompigbrainbyBarderetal.[1]in1982,belongstothefamilyofneurotrophins(NTs)aswellasnervegrowthfactor(NGF),neurotrophin-3(NT-3),NT-4/5.Itisabletopromotesurvivalanddifferentiationofseveralpopu-lationsofneurons,includingmesencephalicdopaminergicneurons,motorneurons,andcholiner-gicneurons,andtoprotectthemagainstneurotoxicityandischemia.BDNFplaysanimportantroleinregulatingneuronsurvivalanddifferentiationduringdevelopmentandinmaintainingthe…     

从噬菌体展示的构象限制性随机肽库中筛选有 trkB 结合活性的小肽

脑源性神经营养因子(brain-derived neurotrophic factor, BDNF)能够促进多种神经元的存活和分化, 在神经退行性疾病的治疗中具有广阔的应用前景, 但由于其分子量大而无法穿过血脑屏障, 限制了其临床应用. 以表达于NIH 3T3细胞上的BDNF受体trkB为靶分子, 从噬菌体展示的随机肽库中筛选与trkB有亲和力的小肽, 采用不表达trkB 的NIH 3T3细胞预吸附和最后一轮筛选以1 μg/mL的BDNF竞争性洗脱策略, 使能与trkB天然构象结合、又能有效拮抗BDNF与trkB结合的特异性噬菌体得到了有效富集, 获得了5种trkB结合小肽, 它们有核心序列CRA/TXφXXφXXC(X代表随机残基, φ为T, L或I). 对展示有5种外源小肽的噬菌体进行了初步生物活性测定, 未发现有BDNF样活性, 反而能剂量依赖性地拮抗BDNF的生物功能. 依据其中一个克隆序列(C1)合成的肽也证实有拮抗BDNF的作用, 表明获得的5种小肽可能为trkB受体拮抗剂. 这些trkB拮抗性小肽在治疗神经母细胞瘤和慢性痛以及神经生物学研究中具有应用前景, 同时也为构建二级噬菌体展示肽库、继续筛选BDNF模拟小肽奠定了基础.     

庚型肝炎病毒转基因小鼠的建立

利用受精卵原核显微注射的方法,产生了含有HGV结构蛋白C、E1、E2及部分非结构蛋白NS2、NS3的转基因小鼠.得到10只founder小鼠,其中有3只founder小鼠与正常小鼠交配后得到了整合有外源基因的F1代阳性小鼠.RT-PCR的结果显示,外源基因可在founder小鼠及F1代小鼠的血液有核细胞及肝细胞内转录;组织病理学检查显示,某些转基因小鼠的肝细胞出现了水样变、脂肪变性及轻微炎性反应等病理学改变,但与同一品系的正常小鼠相比,转基因小鼠血清转氨酶无明显升高.