棕色固氮菌突变株DJ35 中钼铁蛋白和HBP59 蛋白的研究—纯化特性及晶体生长

我们从A. vinelandii突变株DJ35中纯化得到了ΔnifE Av1，并通过与OP Av1相比较对其特性进行了研究。虽然ΔnifE Av1以四聚体形式(α2β2)存在，但却表现出两种天然电泳行为。与OP Av1相比，ΔnifE Av1中的金属含量较低，每个蛋白分子仅含9.96个铁原子、0.36个钼原子。OP Av1和ΔnifE Av1的吸收光谱相似，均在280 nm附近出现蛋白吸收峰，在300-600 nm波长范围内光吸收普遍降低，并无特征峰出现。虽然ΔnifE Av1可见光区域CD谱的摩尔消光系数（Δε）总比OP Av1小，但在520nm区域附近摩尔消光系数减少的相对值明显大于在450nm附近摩尔消光系数减少的相对值。ΔnifE Av1的EPR信号明显与OP Av1不同，在g≈3.7处的EPR信号完全消失，在g≈4.3 和2.0处的EPR信号强度也分别降低了75%和50%以上。ΔnifE Av1不能与Fe蛋白组成活性单位，但被FeMoco激活后可与Fe蛋白组成活性单位。上述结果表明, ΔnifE Av1不含FeMoco，但具有与OP Av1相同的P-cluster。 我们在纯化ΔnifE Av1的过程中，发现一个类似OP Av1的β亚基的污染蛋白，经MALDI-TOF质谱鉴定这种蛋白是棕色固氮菌中一预测基因的产物。通过改进纯化方法, 我们首次得到80%电泳纯的蛋白，并将其命名为HBP59蛋白。HBP59为一单体蛋白，分子量约为59k Da。金属测定结果表明每个蛋白分子中含有0.42个铁原子。HBP59蛋白的可见光谱表现出典型的血红素蛋白光谱特征，还原态的HBP59蛋白在421nm处有最大吸收峰，同时在517nm、556nm处有两个伴随肩峰出现, A421/A280仅为0.146。HBP59蛋白氧化后，吸收峰发生蓝移，最大吸收峰从421nm移到413nm，517nm和556nm处的肩峰消失，A413/A280为0.168。HBP59蛋白的可见光区圆二色谱比较复杂，正峰出现在420nm, 406nm, 379nm 和364nm; 其摩尔消光系数分别为0.75, 0.94, 0.68 和 0.99;负峰出现在433 nm 和392nm，其摩尔消光系数分别为-1.13 和-0.78。血红素滴定实验结果说明每个HBP59蛋白分子结合了0.1个血红素，但每个蛋白分子具有最大结合1个血红素的能力。这低结合能力与这低比率的比率是非常一致的。该蛋白对温度相对敏感，高于40℃蛋白开始沉淀。上述结果说明HBP59是一个结合具有低自旋和不对称的血红素的蛋白，它在菌体内可能并未扮演贮藏血红素的角色，而是可能参与血红素的运输或附着。 此外，经过结晶条件的大量筛选后获得了ΔnifE Av1和ΔnifH Av1的优质小单晶,并对HBP59的晶体培养进行了初步探索。

A micromechanical model of influence of particle fracture and particle cluster on mechanical properties of metal matrix composites

A general analytical model for a composite with an isotropic matrix and two populations of spherical inclusions is proposed. The method is based on the second order moment of stress for evaluating the homogenised effective stress in the matrix and on the secant moduli concept for the plastic deformation. With Webull's statistical law for the strength of SiCp particles, the model can quantitatively predict the influence of particle fracture on the mechanical properties of PMMCs. Application of the proposed model to the particle cluster shows that the particle cluster has neglected influence on the strain and stress curves of the composite. (C) 1998 Elsevier Science B.V.

Impact of carrier stiffness and microtopology on two-dimensional kinetics of P-selectin and P-selectin glycoprotein ligand-1 (PSGL-1) interactions

Mechanics and surface microtopology of the molecular carrier influence cell adhesion, but the mechanisms underlying these effects are not well understood. We used a micropipette adhesion frequency assay to quantify how the carrier stiffness and microtopology affected two-dimensional kinetics of interacting adhesion molecules on two apposing surfaces. Interactions of P-selectin with P-selectin glycoprotein ligand-1 (PSGL-1) were used to demonstrate such effects by presenting the molecules on three carrier systems: human red blood cells (RBCs), human promyelocytic leukemia HL-60 cells, and polystyrene beads. Stiffening the carrier alone or in cooperation with roughing the surface lowered the two-dimensional affinity of interacting molecules by reducing the forward rate but not the reverse rate, whereas softening the carrier and roughing the surface had opposing effects in affecting two-dimensional kinetics. In contrast, the soluble antibody bound with similar three-dimensional affinity to surface-anchored P-selectin or PSGL-1 constructs regardless of carrier stiffness and microtopology. These results demonstrate that the carrier stiffness and microtopology of a receptor influences its rate of encountering and binding a surface ligand but does not subsequently affect the stability of binding. This provides new insights into understanding the rolling and tethering mechanism of leukocytes onto endothelium in both physiological and pathological processes.

Quantitative structure-property relationships for octanol-air partition coefficients of polychlorinated naphthalenes, chlorobenzenes and p,p '-DDT

The octanol-air partition coefficient (K-OA) is a key descriptor of chemicals partitioning between the atmosphere and environmental organic phases. Quantitative structure-property relationships (QSPR) are necessary to model and predict KOA from molecular structures. Based on 12 quantum chemical descriptors computed by the PM3 Hamiltonian, using partial least squares (PLS) analysis, a QSPR model for logarithms of K-OA to base 10 (log K-OA) for polychlorinated naphthalenes (PCNs), chlorobenzenes and p,p'-DDT was obtained. The cross-validated Q(cum)(2) value of the model is 0.973, indicating a good predictive ability of the model. The main factors governing log K-OA of the PCNs, chlorobenzenes, and p,p'-DDT are, in order of decreasing importance, molecular size and molecular ability of donating/accepting electrons to participate in intermolecular interactions. The intermolecular dispersive interactions play a leading role in governing log K-OA. The more chlorines in PCN and chlorobenzene molecules, the greater the log K-OA values. Increasing E-LUMO (the energy of the lowest unoccupied molecular orbital) of the molecules leads to decreasing log K-OA values, implying possible intermolecular interactions between the molecules under study and octanol molecules. (C) 2002 Elsevier Science Ltd. All rights reserved.

Vertical variations in kinetics of alkaline phosphatase and P species in sediments of a shallow Chinese eutrophic lake (Lake Donghu)

The vertical distribution of the variables relevant to P forms in sediments were studied in a shallow Chinese freshwater lake (Lake Donghu) in 1997, 1998, 1999 and 2000, to assess the contribution of enzyme to P availability in sediment cores. Sediment P was fractionationd into iron-bound P, calcium-bound P, acid soluble organic P (ASOP) and hot NaOH extractable residual organic P. The former two species made the largest contribution to the sediment P pool. All P species exhibited significantly higher concentrations in different depths at Station I, compared with those found at Station II, except for ASOP. Coupled with these lower ASOP concentrations, the V-max data of alkaline phosphatase, measured on the same samples, were significantly higher at station I. Taken together, ASOP were probably important in supplying the enzymatic substrate (Phosphatase Hydrolyzable Phosphorus, PHP) into interstitial water. Dissolved orthophosphate and PHP concentrations were highly heterogeneous , but peaked in subsurface, paralleled by higher V-max and lower K-m values of alkaline phosphatase, throughout the sediment core. Sediment in the eutrophic lake is not only enriched in available P (iron-bound P), or stores residual P, but also tends to release PHP, thereby inducing the production of alkaline phosphatase and releasing o-P into water column by enzymatic hydrolysis. The latter process may also occur in relatively deep sediment layers.

Electronic structures of wurtzite ZnO, BeO, MgO and p-type doping in Zn1-xYxO (Y = Mg, Be)

Using the density function theory within the generalized gradient approximation, the band structures of wurtzite ZnO, BeO and MgO have been calculated. The effective-mass parameters are fitted using the calculated eigenvalues. The Dresselhaus spin-orbit effect appears in the k[1 00] direction, and is zero in the high symmetry direction k[00 1]. The orderings of valence band split by the crystal-field and spin-orbit coupling in wurtzite ZnO, BeO and MgO are identified by analyzing the wave function characters calculated by projecting the wave functions onto p-state in the spherical harmonics. For wurtzite ZnO, the ordering of valence band is Still Gamma(7) > Gamma(9) > Gamma(7) due to the negative spin-orbit coupling splitting energy and the positive crystal-field splitting energy. Thus, the Thomas' conclusion is confirmed. For wurtzite BeO and MgO, although their orderings of valence bands are Gamma(7) > Gamma(9) > Gamma(7) too, the origins of their orderings are different from that of wurtzite ZnO. Zn1-x,YxO (Y = Mg, Be) doped with N and P atoms have been studied using first-principles method. The calculated results show that N atom doped in Zn1-x BexO has more shallow acceptor energy level with increasing the concentration of Be atom. (C) 2008 Elsevier B.V. All rights reserved.

Ultrafast carrier dynamics in undoped and p-doped InAs/GaAs quantum dots characterized by pump-probe reflection measurements

We investigate the dependence of the differential reflection on the structure parameters of quantum dot (QD) heterostructures in pump-probe reflection measurements by both numerical simulations based on the finite-difference time-domain technique and theoretical calculations based on the theory of dielectric films. It is revealed that the value and sign of the differential reflection strongly depend on the thickness of the cap layer and the QD layer. In addition, a comparison between the carrier dynamics in undoped and p-doped InAs/GaAs QDs is carried out by pump-probe reflection measurements. The carrier capture time from the GaAs barrier into the InAs wetting layer and that from the InAs wetting layer into the InAs QDs are extracted by appropriately fitting differential reflection spectra. Moreover, the dependence of the carrier dynamics on the injected carrier density is identified. A detailed analysis of the carrier dynamics in the undoped and p-doped QDs based on the differential reflection spectra is presented, and its difference with that derived from the time-resolved photoluminescence is discussed. (C) 2008 American Institute of Physics.