一种新的(1→3)-β-D-葡聚糖识别蛋白的鉴定及其分子克隆

目的从大黄粉虫 (Tenebriomolitor)体内鉴定出一种 (1→ 3) β D 葡聚糖识别蛋白 (Tm GRP)并对其进行分子克隆。方法通过制备一种 β 1,3 葡聚糖固定化柱 ,从大黄粉虫的体液中筛选出能与 (1→ 3) β D 葡聚糖发生特异性结合的蛋白质。利用DNA探针法和随机引物法从大黄粉虫的cDNA文库中筛选Tm GRP的全部DNA序列。结果Tm GRP是一个由 14 4 3个核苷酸编码产生的具有 4 81个氨基酸残基的蛋白质。结论Tm GRP是大黄粉虫体内的一种葡聚糖识别蛋白。     

苯硫脲嗪对棉鼠丝虫和马来丝虫感染的治疗

新化合物苯硫脲嗪(4-硝基-4’-(1-甲基哌嗪-4-硫代甲酰氨基)二苯胺)具有抗丝虫作用。感染棉鼠丝虫的棉鼠或长爪沙鼠和感染马来丝虫的长爪沙鼠用苯硫脲嗪ig 200mg/(kg·d)×3d皆获治愈。苯硫脲嗪对微丝蚴亦有作用,给药开始后3d,受治棉鼠周围血中的微丝蚴即消失,而受治长爪沙鼠腹腔内的马来丝虫微丝蚴活动减弱、僵直或自溶。两种丝虫经苯硫脲嗪作用后,虫体被破坏、崩解,特别是马来丝虫,出现大量死虫肉芽肿,虫周细胞反应强烈。     

呼吸道合胞病毒模拟表位合成肽疫苗的开发

开发诱导呼吸道合胞病毒 (RSV)中和抗体的合成肽疫苗是预防 RSV感染的有效途径。本文叙述了用针对 RSV F蛋白构象表位的单克隆抗体 MAb1 9筛选固相组合肽库 ,鉴定出与MAb1 9发生特异性反应的两个序列 ,并经氨基酸替换试验 ,提高了与 MAb1 9反应的亲和力。将模拟表位以多抗原肽 (MAP)形式免疫 Balb/ c小鼠 ,表现出明显的保护作用。     

呼吸道合胞病毒模拟表位合成肽疫苗的开发

并发诱导呼吸道合胞病毒（RSV）中和抗体的合成肽疫苗是预防RSV感染的有效途径。本文叙述了用针对RSV F蛋白构象表位的单克隆抗体MAb19筛选固组组合肽库，鉴定出与MAb19发生特异性反应的两个序列，并经氨基酸替换试验，提高了与MAb19反应的亲和力，将模拟表位以多抗原位（MAP）形式免疫Balb/c小鼠，表现出明显的保护作用。     

蛋白质合成抑制剂的筛选测定

蛋白质的合成是目前临床上应用的抗菌剂的有效靶位.许多已知的蛋白质合成抑制剂(四环素类、氨基糖苷类、大环内酯类等)都是天然产物.涉及到蛋白质合成独特反应的分子上的和机制上的细节尚未被完全阐明,因此出现了理性化设计的抑制剂.     

B型肉毒毒素重链C端基因的克隆与表达

目的:扩增B型肉毒毒素重链C端基因并将其在大肠杆菌中表达.方法:首先克隆B型肉毒毒素重链C端片段(BoNTB/Hc),经IPTG诱导,在大肠杆菌中进行天然序列的表达.构建原核表达载体pET32/Hc后进行融合蛋白的表达.对5′端引物进行修饰,最终进行N端修饰蛋白的表达.结果:扩增得到的BoNTB/Hc与已知序列同源性达99%,未得到天然序列的表达,获得了融合蛋白和N端修饰蛋白的表达.Western blot鉴定结果表明,融合蛋白和N端修饰蛋白都可以和特异性抗体发生反应.结论:获得了B型肉毒毒素重链C端基因的表达,为肉毒毒素相关研究奠定了基础.     

B型肉毒毒素重链C端基因的克隆与表达

目的:扩增B型肉毒毒素重链C端基因并将其在大肠杆菌中表达.方法:首先克隆B型肉毒毒素重链C端片段(BoNTB/Hc),经IPTG诱导,在大肠杆菌中进行天然序列的表达.构建原核表达载体pET32/Hc后进行融合蛋白的表达.对5′端引物进行修饰,最终进行N端修饰蛋白的表达.结果:扩增得到的BoNTB/Hc与已知序列同源性达99%,未得到天然序列的表达,获得了融合蛋白和N端修饰蛋白的表达.Western blot鉴定结果表明,融合蛋白和N端修饰蛋白都可以和特异性抗体发生反应.结论:获得了B型肉毒毒素重链C端基因的表达,为肉毒毒素相关研究奠定了基础.     

抗菌素

124.膦肽,一类新的抗细菌合成药物已知青霉素、头孢菌素、D-环丝氨酸及万古霉素抑制形成细菌细胞壁粘肽的末端D-Ala-D-Ala(D-丙氨酸-D-丙氨酸)或抑制D-Ala-D-Ala与粘肽中二氨基庚二酸的交联。作者于1971年开始研究类似于D-Ala-D-Ala的合成肽。用C-端氨基酸与各种L-及D-α-氨基羧酸合成300多种二肽到五肽类似物。结果发现,某些膦肽,特别是含有L-Ala(p)(L-1-氨基乙基膦酸)残基的化     

具有高度免疫原性的抗C亚型HIV-1合成肽疫苗构建物的制备

一些疾病的治疗已经从合成肽疫苗的使用中获益。作者将两种不同的肽合成方式结合起来制备成一种含多种表位免疫原 ( MEI)的合成肽免疫原 ,并用动物模型测定了这种合成肽疫苗构建物诱导的免疫应答。　　 MEI序列衍生于 C亚型 1型人类免疫缺陷病毒 ( HIV- 1 )包膜的 V3区 ,肽序列内的四聚体端点 GPGQ为型特异性中和抗体的主要结合位点。在合成时 ,按既定百分率和位点混合氨基酸使一些氨基酸偶联 ,这种方法获得的肽混合物代表 6 .7× 1 0 7个包膜序列 ,其中包括已知的 339个序列中的 30 1个序列以及 V3序列。通过疏水图等观察分析 ,证实…     

Differential in vitro association of vinca alkaloid-induced tubulin spiral filaments into aggregated spirals.

Vinblastine, vincristine, vindesine, and vinorelbine, the four vinca alkaloids used in cancer therapy, differ in their antitumoral spectra and toxicities, but not in their inhibitory effects on microtubule assembly in vitro. At higher drug concentrations, vinca alkaloids induce the assembly of spiral filaments of tubulin, which, in turn, can interact laterally and form paracrystals. Using methods that distinguish spiral filaments and paracrystals (aggregated spirals), we found that spiral filament formation was largely independent of the incubation temperature, of the alkaloid used, and of the presence or absence of microtubule-associated proteins (MAPs). In contrast, the formation of aggregated spirals was markedly dependent on the alkaloid used, on the incubation temperature, and on the absence or presence of MAPs. Aggregated spirals failed to assemble in the presence of high concentrations of MAP-1A or MAP-1B, whereas they assembled readily with tau and MAP-2. Differences in patterns of turbidity development using pure tubulin allowed the classification of thirteen cytotoxic vinca alkaloids into five distinct groups, with centrifugal recovery of aggregated spirals in close agreement with the various turbidity patterns. With microtubule protein, i.e. tubulin preparations containing MAPs, only four groups were defined by turbidity patterns, and centrifugal protein recovery was more divergent. Vinblastine, vincristine, vindesine, and vinorelbine fell into distinct groups under both reaction conditions, and thus they appear to have qualitatively distinguishable in vitro interactions with tubulin. These differential effects on spiral filament and aggregated spiral assembly revealed that the four drugs induce different constraints on the tubulin molecule.     

rhG-CSF增加实验性脑缺血周围区微管相关蛋白的表达

目的研究重组人粒细胞集落刺激因子(Reeomhinant human gmnulocyte colony-stimulating factor,rhG-CSF)对脑梗死周围区的微管相关蛋白(Microtubule-associated protein 2,MAP-2)表达的影响。方法用线栓法建立大鼠大脑中动脉缺血再灌注模型,观察rhG-CSF对神经功能缺损评分的影响；应用半定量逆转录一聚合酶链式反应(RT-PCR)方法和免疫组织化学方法分别测定梗死周围区MAP-2的mRNA和蛋白表达水平的变化。结果与对照组相比,治疗组的神经功能缺损评分较低,脑缺血周围区MAP-2的mRNA和蛋白表达增高。结论rhG-CSF可以促进脑缺血周围区的微管相关蛋白的表达,从而减轻脑缺血再灌注损伤。     

Proteomic analysis of lysosomal proteins in melanocyte B16F10 exposed to melanin

A high quantity of melanin causes hyperpigmentation or skin cancer in humans and it can also activate lysosomal enzymes much melanin. In this study, lysosomal proteins were identified through two-dimensional polyacrylamide gel electrophoresis (2-DE) after melanocytes were exposed to melanin (B16F10). Melanocytes were exposed to 100 ppm melanin for 24 hours and the lysosomal proteins were extracted. The results showed that 18 protein spots up regulated and 6 protein spots down regulated after melanin exposure. The expression pattern of established lysosomal proteins is informative in understanding proteins related to decreasing melanin.     

Zinc stimulation of bone protein synthesis in tissue culture. Activation of aminoacyl-tRNA synthetase

The present investigation was undertaken to clarify the effect of zinc on bone protein synthesis in tissue culture. Calvaria were removed from 3-week-old male rats and cultured for periods up to 96 hr in Dulbecco's Modified Eagle Medium (high glucose, 4500 mg/dl) supplemented with antibiotics and bovine serum albumin. The calvaria were incubated at 37 degrees in 5% CO2/95% air in the medium containing 10(-6)-10(-4) M zinc. Zinc content in bone cells was increased when the culture was treated with 10(-5) and 10(-4) M zinc for 48 hr. When calvaria cultured in the presence of 10(-4) M zinc were pulsed with [14C]uridine, the incorporation of [14C]uridine into the bone RNA was not increased significantly. In the pulse with [3H]leucine, the presence of 10(-5) to 10(-4) M zinc in the medium caused a significant increase in the incorporation of [3H]leucine into the acid-insoluble residues of bone tissue. This increase was blocked completely by treatment with 10(-7) M cycloheximide, an inhibitor of protein synthesis. When [3H]leucine was added into the reaction mixture containing the 5500 g supernatant fraction of the homogenate prepared from calvaria cultured in the presence of 10(-4) M zinc, the in vitro protein synthesis was increased about 2-fold. The activity of [3H]leucyl-tRNA synthetase in the 105,000 g supernatant fraction (cytosol) of the bone homogenate was increased about 2-fold by the culture with 10(-4) M zinc. The presence of 10(-4) M dipicolinate, a specific chelator of zinc, in the culture medium negated the effect of zinc on [3H]leucyl-tRNA synthetase activity. The addition of 10(-7) to 10(-6) M zinc into the reaction mixture containing enzyme extracts obtained from uncultured rat calvaria caused a 2-fold increase of [3H]leucyl-tRNA synthetase activity. These results clearly indicate that zinc induces the stimulation of protein synthesis at the translational level in bone cells. The present study further supports the view that zinc increases protein synthesis in bone cells and that the metal induces bone formation.     

Phloridzin-induced melanogenesis is mediated by the cAMP signaling pathway

Melanogenesis is a physiological process that results in the synthesis of melanin pigments, which play a crucial protective role in protection against skin photocarcinogenesis. Phloridzin is a phloretin 2′-glucoside that is found in many parts of the apple tree that reportedly increases tyrosinase activity and melanin contents through inhibition of protein kinase C (PKC) activity in B16 melanoma cells. In this study, we attempted to accurately determine the effects and mechanisms of action of phloridzin on melanogenesis. Specifically, we observed that phloridzin-induced a dose-dependent increase in tyrosinase activity and melanin contents, and that these changes were accompanied by an increase in the levels of tyrosinase and the tyrosinase-related proteins, TRP-1 and TRP-2. Furthermore, the cAMP-dependent protein kinase A (PKA) inhibitor H89 impaired the response of the tyrosinase activity and melanin synthesis to phloridzin. Additionally, phloridzin stimulated cAMP production and phosphorylation of the cAMP-response element binding protein (CREB). Taken together, the results of this study indicate that phloridzin increases tyrosinase gene expression through the cAMP signaling pathway, thereby leading to the stimulation of melanogenesis.     

Lysosomal membrane proteins exposed to melanin in melanocyte B16F10

There have been studied on therapeutic of anticancer and antimicrobial using by effective proteins or peptides. In this study, we examined the lysosomal membrane protein extracted from lysosomes isolated from melanocyte treated by melanin with 100 ppm for 24 hours in order to find them reacted to targeting ability to melanin using by 2-dimensional electrophoresis (2DE). Consequently, we found 29 up-regulated (2.0 fold) and 23 down-regulated (2.0 fold) spots associated with melanin treatment. This result shown that lysosomal membrane proteins have function to react to treatment or decrease of melanin quantity through cellular activation of lysosomes.