应用免疫斑点法检测不同工艺灭活的乙肝血源疫苗中的前S_1和S_2蛋白

作者应用前 S_1、前 S_2和 HBsAg/a 单克隆抗体,用免疫斑点法(Immuno-spot)检测同一批的乙肝表面抗原分别经加热灭活和三步化学灭活后的前 S_1和前 S_2蛋白保留情况,比较了两种工艺对前 S_1和前 S_2蛋白的影响。结果表明;加热灭活可保留前 S_1和前 S_2蛋白,三步化学灭活使前 S_1和前 S_2蛋白丢失,从抗原组成上看,加热灭活后的抗原更接近自然抗原。首次报告了含有前 S_1蛋白的乙肝疫苗,并对前 S_1和前 S_2蛋白在乙肝血源疫苗中的可能作用进行了讨论。     

Protein design to understand peptide ligand recognition by tetratricopeptide repeat proteins

Protein design aims to understand the fundamentals of protein structure by creating novel proteins with pre-specified folds. An equally important goal is to understand protein function by creating novel proteins with pre-specified activities. Here we describe the design and characterization of a tetratricopeptide (TPR) protein, which binds to the C-terminal peptide of the eukaryotic chaperone Hsp90. The design emphasizes the importance of both direct, short-range protein-peptide interactions and of long-range electrostatic optimization. We demonstrate that the designed protein binds specifically to the desired peptide and discriminates between it and the similar C-terminal peptide of Hsp70.     

Comparison of the construction of a 3-D model for human thromboxane synthase using P450cam and BM-3 as templates: implications for the substrate binding pocket

A 3-D model of human thromboxane A₂ synthase (TXAS) was constructedusing a homology modeling approach based on information fromthe 2.0 crystal structure of the hemoprotein domains of cytochromeP450BM-3 and P450cam. P450BM-3 is a bacterial fatty acid monooxygenaseresembling eukaryotic microsomal cytochrome P450s in primarystructure and function. TXAS shares 26.4% residue identity and48.4% residue similarity with the P450BM-3 hemoprotein domain.The homology score between TXAS and P450BM-3 is much higherthan that between TXAS and P450cam. Alignment between TXAS andthe P450BM-3 hemoprotein domain or P450cam was determined throughsequence searches. The P450BM-3 or P450cam main-chain coordinateswere spplied to the TXAS main chain in those sements where thetwo sequences were well aligned. These segments were linkedto one another using a fragment search method, and the sidechains were added to produce a 3-D model for TXAS. A TXAS substrate,prostaglandin H₂ (PGH₂) was docked into the TXAS cavity correspondingto the arachidonic acid binding pocket in P450BM-3 or camphorbinding site in P450cam. Regions of the heme and putative PGH2binding cavities in the TXAS model were identified and analyzed.The segments and residues involved in the active-site pocketof the TXAS model provide reasonable candidates for TXAS proteinengineering and inhibitor design. Comparison of the TXAS modelbased on P450BM-3 with another TXAS model based on the P450BM-3with another TXAS model based on the P450cam structure indicatedthat P450BM-3 is a more suitable template for homology modelingof TXAS.     

Preparation and surface activity of modified soy protein

A series of novel surfactants have been prepared by the reaction of hydrolyzed soy protein with alkyl succinic anhydride. These novel surfactants exhibit excellent surface active properties including surface tension, foaming, emulsifying, wetting power, and buffer ability. The hydrophobic modified protein exhibit more surface activity than original protein because of the enhanced hydrophobicity in structure. The increase in hydrophobic chain length leads to an increase in the surface activity. The surface tension reduction is correlated to the hydrophobicity of the modified molecule, which was determined by a fluorescent probe. In application on cotton bleaching procedures, these surfactants increase the whiteness of fabrics. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 102: 3498–3503, 2006     

应用骨形成蛋白与胶原膜复合物修复大鼠颅骨缺损

目的探讨骨形成蛋白(BMP)与胶原膜复合物(复合膜)修复大鼠颅骨缺损的效果。方法将BMP与胶原膜复合,在大鼠颅骨制备骨缺损,分别采用双侧覆盖复合膜、外侧覆盖复合膜进行治疗,以外侧覆盖胶原膜作为平行对照,并设空白对照。于术后2、4、6周,取标本进行X线检查及组织学观察。结果 两个覆盖复合膜组的缺损成骨面积百分比明显高于覆盖胶原膜的平行对照组(P<0.01)。术后4、6周,双侧覆盖复合膜组的成骨面积百分比明显大于单侧覆盖复合膜组(P<0.05)。双侧覆盖复合膜组的缺损术6周后已达骨性愈合。结论BMP与胶原膜复合既具有机械性阻挡作用,又具有骨诱导性,能加速骨缺损愈合。     

Tissue-type plasminogen activator variants with domain duplications and rearrangements

The interactions between tPA domains that are important forcatalysis are poorly understood. We have probed the functionof interdomain interactions by generating tPA variants in whichdomains are duplicated or rearranged. The proteins were expressedin a transient mammalian expression system and tested in vitrofor their ability to activate plasminogen, induce fibrinolysisand bind to a forming fibrin clot. Duplication of the heavychain domains of tPA produced enzymatically active tPA variants,many of which demonstrated similar in vitro amidolytic and fibrinolyticactivity and similar fibrin affinity to the parent molecule.Zymographic analysis of the domain duplication tPA variantsshowed one major active species for each variant. Selectionof the residues duplicated and the interdomain spacing werefound to be critical considerations in the design of tPA variantswith duplicated domains. We also rearranged the domains of tPAsuch that kringle 1 replaced the second kringle domain and viceversa. An analysis of these variants indicates that the firstkringle domain can confer fibrin affinity to a tPA variant andfunction in place of kringle 2. Therefore, in wild-type tPA,the functions of kringle 1 and kringle 2 must be dependent partiallyon their orientation within the heavy chain of the protein.The functional autonomy of the heavy and light chains of tPAis demonstrated by the activity of a tPA variant in which theorder of the heavy and light chains was reversed.     

Free energy calculations of the mutation of Ile96 -> Ala in barnase: contributions to the difference in stability

Free energy calculations were carried out to determine the relativeunfolding free energy of the Ile96 wild type and Ala96 mutantbarnases. The total calculated free energies suggest that substitutionof Ile96 with Ala destabilizes barnase by 3.9 kcal/mol, whichis in good agreement with the independently determined experimentalvalues of 4.0 and 3.3 kcal/mol and a previous simulation. However,a decomposition of the free energy finds the dominant contributionsto this free energy arising from the noncovalent Interactionsbetween the perturbed group and distant residues of barnasein the sequence and water molecules and only a very small contributionfrom covalent interactions. This is in contrast to the previoussimulation, using the dual topology methodology, which produceda decomposition with an {small tilde}60% free energy contributionfrom changes in covalent interactions. The use of the singletopology employed in the present calculations and the dual topologyemployed in the previous study are analyzed in order to understandthe contrast between the present results and the results ofthe previous study.     

中空纤维超滤膜去除乳酸钙中蛋白质的研究

发酵法联合生产乳酸-乳酸钙工艺中,分离出的乳酸钙产品含有蛋白质、还原糖、重金属等杂质,影响产品的品质。本实验研究了中空纤维膜对乳酸钙中蛋白质的去除,并分析了压差、温度、浓度及pH值等因素的影响。得到了较好的工艺操作条件为Δp=0.05MPa,Δ=30g/L,30°C,pH=4。进一步提出用多步超滤能提高蛋白质的去除率15%～27%。     

中国流行株HIV-1B/C重组亚型包膜蛋白真核表达载体的构建及稳定表达细胞株的筛选

目的获得高质量的中国流行株HIV-1B/C重组亚型包膜蛋白抗原。方法改造表达载体,以构建中国流行株HIV-1B/C重组亚型包膜蛋白基因带有筛选标记的真核细胞表达质粒,将所构建质粒转染真核细胞,用含有抗性的培养基筛选出能够高效、持续表达包膜蛋白的稳定表达细胞株。结果所构建的表达载体pVRPEnv转染细胞后可表达目的蛋白,并建立了稳定表达细胞系。结论获得了可以高效、持续稳定表达中国流行株HIV-1B/C重组亚型包膜蛋白的细胞株。