A “Three-in-One” Multifunctional Probe for Bcl-2/Mcl-1 Profiling and Visualizing in Situ

Bcl-2 and Mcl-1, the two arms of the anti-apoptotic Bcl-2 family proteins, have been identified as key regulators of apoptosis and effective therapeutic targets of cancer. However, no small-molecular probe is capable of profiling and visualizing both Bcl-2 and Mcl-1 simultaneously in situ. Herein, we report a multifunctional molecular probe (BnN₃-OPD-Alk) by a “three-in-one” molecular designing strategy, which integrated the Bcl-2/Mcl-1 binding ligand, fluorescent reporter group and photoreactive group azido into the same scaffold. BnN₃-OPD-Alk exhibited sub-micromolar affinities to Bcl-2/Mcl-1 and bright green self-fluorescence. It was then successfully applied for Bcl-2/Mcl-1 labeling, capturing, enriching, and bioimaging both in vitro and in cells. This strategy could facilitate the precise early diagnosis and effective therapy of dual Bcl-2/Mcl-1-related diseases.     

The mechanical property and phase structures of wheat proteins/polyvinyl alcohol blends studied by high-resolution solid-state NMR

The phase structures of thermally processed wheat proteins (WP) and polyvinyl alcohol (PVOH) blends were studied by solid-state high-resolution NMR spectroscopy. The intermolecular interactions among the multi-component systems and the behavior of each component in the blends on scales of nanometers were examined. The mechanical properties of the blends were also measured and related to the phase structure studies. The results indicated that the polymer chains of WP could be homogeneously mobilized when thermally processed with glycerol and water as plasticisers, but the glycerol predominately associated with WP rather than PVOH in the blends. The intermolecular hydrogen bonding interactions between WP and PVOH caused some extent of miscibility in the system on scales of nanometers especially when the PVOH content was low. The tensile strength and modulus of the blends were improved as compared to WP. However, the intermolecular interactions were relatively weak and could not be further enhanced by increasing PVOH component in the blends. The particle miscible WP/PVOH blends contained plasticised WP and PVOH phases in conjunction with the miscible WP/PVOH phase. Increasing the PVOH content in the blends did not result in an increase of the percentage of the miscible phase and the blends tented to be immiscible while the elongation of the blends was reduced when increasing the PVOH content in the blends.     

STUDIES ON COUNTERACTING CHROMATOGRAPHIC ELECTROPHORESIS AND ITS UTILIZATION FOR PROTEIN PURIFICATION

A counteracting chromatographic electrophoresis(CACE)separation device has been success-fully built and tested.Experiments are performed with protein mixtures of BSA,Hg,Mg and Cyto-cto study the separation performances,concentration profiles,start-up methodology as well as influ-ences of pH and ionic strength on the separation process.Mathematical models based on the fluiddynamics of the porous medium are proposed.Experiments indicate that the CACE process has agood commercial potential in protein engineering and fine chemicals purification.     

蛋白质增敏钼-邻苯二酚紫体系的研究及应用

在pH=2.8的clark-lubs缓冲介质中,牛血清白蛋白能显著增敏钼-邻苯二酚紫的显色体系,据此建立了分光光度法测定钼的新方法。仔细考察了反应酸度、邻苯二酚紫浓度、牛血清白蛋白浓度及反应时间等因素对测定的影响。最大吸收波长在680 nm处,与钼-邻苯二酚紫体系相比,红移了120 nm。10 mL溶液中,钼质量在0～40μg范围内服从比尔定律,测定的表观摩尔吸光系数为2.11×104L/(mol.cm)。所拟方法简便、快速、灵敏度高,用于合金钢样品中微量钼的测定,结果与认定值一致。     

黄粉虫纤溶活性蛋白免疫原性与抗血栓作用的研究

为了研究黄粉虫纤溶活性蛋白（ Tenebrio Fibrinolytic Proteins,TFP）的免疫原性及其抗血栓作用,本研究将纯化的TFP经皮下注射免疫小鼠,间接ELISA法检测小鼠血清中的抗体含量及抗体效价;小鼠灌胃TFP后,取血浆检测凝血酶时间（thrombin time,TT）、凝血酶原时间（prothrombin time,PT）、活化部分凝血活酶时间（activated partial thromboplastin time ,APTT）、组织型纤溶酶原激活剂（ tissue-type plasminogen activator ,t-PA）和其抑制物（ plasminogen activator inhibitor ,PAI）,角叉菜胶使小鼠尾部形成血栓,测量血栓形成长度。结果显示,TFP免疫小鼠后产生特异性抗体且随着免疫次数的增加,抗体含量逐渐升高,末次免疫后产生的抗体效价为1∶640;TFP可以显著延长TT,增强t-PA的活性,抑制PAI的活性,同时显著抑制由角叉菜胶引起的小鼠尾部血栓的形成。这说明TFP具有良好的免疫原性且具有一定的抗血栓作用。     

Bi-layer gelatin active films with “Pitanga” leaf hydroethanolic extract and/or natamycin in the second layer

Research on biopolymers-based active films produced with natural antioxidants and/or antimicrobials has gained attention over the last few years; however, anti-mold activity has been less studied than those of anti-bacteria. The aim of this work was the development and characterization of bi-layer films based on gelatin with natamycin and/or “Pitanga” leaf hydroethanolic extract in the second thin layer in order to determine the effects of these bioactive compounds on bi-layered film properties. The films were characterized regarding their moisture content and solubility in water, optical properties, microstructure, mechanical and thermal properties, water contact angle, water vapor permeability, UV/visible light transmission, X-ray diffraction, Fourier transform infrared (FTIR) spectroscopy, and antioxidant and anti-mold activities. Active films presented activity against Penicillium spp and Aspergillus niger and demonstrated antioxidant activity, as measured by ABTS ^•+ and DPPH ^• methods. Neither additive used in the films' second layer significantly affected the films' moisture content, thermal properties or the molecular interactions of the polymer matrix, assessed by FTIR, although some mechanical properties were affected, and the water contact angle. In conclusion, bi-layer films have reduced the quantity of additives required to maintain the antioxidant and anti-mold activities, as compared to similar monolayer films of the same thickness.     

Continuum electrostatic approach for evaluating positions and interactions of proteins in a bilayer membrane

Orientations of proteins in the membranes are crucial to their function and stability. Unfortunately the exact positions of these proteins in the lipid bilayer are mostly undetermined. Here, the spatial orientation of membrane proteins within the lipid membrane was evaluated using a Poisson–Boltzmann solvent continuum approach to calculate the electrostatic free energy of the protein solvation at various orientations in an implicit bilayer. The solvation energy was obtained by computing the difference in electrostatic energies of the protein in water and in lipid/water environments, treating each as an implicit solvent model. The optimal position of transmembrane proteins (TMP) in a lipid bilayer is identified by the minimum in the “downhill” pathway of the solvation energy landscape. The energy landscape pattern was considerably conserved in various TMP classes. Evaluation of the position of 1060 membrane proteins from the orientations of proteins in membranes (OPM) database revealed that most of the polytopic and β-barrel proteins were in good agreement with those of the OPM database. The study provides a useful scheme for estimating the membrane solvation energy made by lipid-exposed amino acids in membrane proteins. In addition, our results tested with the bacterial potassium channel model demonstrated the potential usefulness of the approach in assessing the quality of membrane protein models. The present approach should be applicable for constructing transmembrane proteins–lipid configuration suitable for membrane protein simulations and will have utility for the structural modeling of membrane proteins.     

The contribution of proteomics towards deciphering the enigma of Coxiella burnetii

Coxiella burnetii, the causative agent of Q fever, is an intracellular bacterium and a potential weapon for bioterrorism. The widespread throughout the world, zoonosis is manifested clinically as a self-limited febrile illness, as pneumonia (acute Q fever) or as a chronic illness with endocarditis being its major complication. The recent Netherlands Q fever outbreak has driven the bacterium from a relatively cryptic, underappreciated, “niche” microorganism on the sideline of bacteriology, to one of possibly great impact on public health. Advances in the study of this microorganism proceeded slowly, primarily due to the, until recently, obligatory intracellular nature of the pathogen that in its virulent phase I must be manipulated under biosafety level-3 conditions. Proteomic studies, in particular, have generated a vast amount of information concerning several aspects of the bacterium such as virulence factors, detection/diagnostic and immunogenic biomarkers, inter-/intraspecies variation, resistance to antibiotics, and secreted effector proteins with significant clinical impact. The phenomenon observed following the genomics era, that of generation and accumulation of huge amount of data that ultimately end up unexploited on several databases, begins to emerge in the proteomics field as well. This review will focus on the advances in the field of C. burnetii proteomics through MS, attempting in parallel to utilize some of the proteomics findings by suggesting future directions for the improvement of Q fever diagnosis and therapy.