犁头尖凝集素色氨酸(Trp)所处微环境及构象的研究

本文通过荧光淬灭的方法对犁头尖凝集素(TDL)的内源荧光进行了淬灭研究.研究结果表明:丙烯酰胺与TDL分子中Trp残基的可接近程度达到100%,而KT和CsCl与Trp残基的可接近程度分别为83.3%和50%,说明TDL中大部分Trp残基位于分子表面或近表面区域,且位于表面的Trp残基所处微环境中带正电荷基团约为带负电荷基团的一倍.荧光淬灭数据分析表明这三种淬灭剂对TDL的荧光淬灭作用均属动态淬灭机制.此外,本文还研究了变性剂对TDL凝血活性与荧光光谱的影响,研究表明同浓度的盐酸胍对TDL变性作用明显强于脲,TDL活性的丧失主要是由于其活性中心被变性剂破坏而造成.     

氨基酸修饰及荧光光谱分析研究黄花石蒜凝集素生物学活性与构象的关系

本文通过对黄花石蒜凝集素(Lycoris aurea Agglutinin,LAA)进行特殊氨基酸的化学修饰,显示一个LAA分子一共有8个色氨酸分子,其中有3个位于分子表面或近表面,Trp、Tyr和Ser/Thr不是LAA凝集活性所必需的氨基酸,而Asp/Glu的羧基和凝血活性密切相关,对其修饰后导致凝血活性丧失50%.通过荧光淬灭的方法对LAA分子中色氨酸所处微环境进行了研究.结果显示中性淬灭剂丙烯酰胺对LAA分子中色氨酸的淬灭作用最强可以淬灭100%的色氨酸荧光,其次是离子型淬灭剂碘化钾,能淬灭62.9%的色氨酸荧光,而氯化铯对LAA色氨酸的淬灭最弱,几乎不能淬灭LAA的荧光.     

家蝇幼虫抗菌肽MDL-3分子结构的荧光特性

研究了家蝇幼虫抗菌肽MDL-3的荧光光谱和淬灭剂对内源性荧光的影响.家蝇幼虫抗菌肽MDL-3在280nm波长的激发光时,荧光光谱为Tyr残基和Trp残基共同提供,KI不能淬灭抗菌肽MDL-3的Trp残基的荧光,而Acr只能淬灭71%(f=0.71)的抗菌肽MDL-3中的Trp残基的荧光,说明Trp残基不是位于抗菌肽分子的表面,而是位于分子的内部.     

可可碱对胰α-淀粉酶的抑制类型及光谱性质的影响

运用紫外光谱和荧光光谱技术研究可可碱与胰α-淀粉酶的结合.反应类型为非竞争性抑制.Ki值为0.72×10-2 mol/L.根据Stern-Volmer方程,计算了可可碱对胰α-淀粉酶的表观淬灭常数,推测其荧光影响可能属于静态淬灭机制.     

咖啡因对胰β-淀粉酶部分理化性质的影响

运用紫外光谱和荧光光谱技术研究咖啡因与胰α-淀粉酶的结合。反应类型为非竞争性抑制。Ki值为0.47×10-2mol/L。根据Stern-Volmer方程,计算了咖啡因对胰α-淀粉酶的表观淬灭常数,推测其荧光影响可能属于静态淬灭机制。     

茶碱与胃蛋白酶相互作用的光谱性质研究

采用紫外光谱法和荧光光谱法研究了茶碱与胃蛋白酶的结合作用。观测到茶碱使胃蛋白酶的紫外吸收峰增强,特征荧光峰淬灭。Stern-Volmer淬灭曲线显示,茶碱对胃蛋白酶的荧光淬灭很可能是一个单一的静态淬灭过程。     

粪臭素对胃蛋白酶部分理化性质的影响

采用紫外光谱法和荧光光谱法研究了粪臭素与胃蛋白酶的结合作用。观测到粪臭素使胃蛋白酶的紫外吸收峰增强,特征荧光峰淬灭。Stern-Volmer淬灭曲线显示,粪臭素对胃蛋白酶的荧光淬灭很可能是一个单一的静态淬灭过程。     

白茯苓凝集素的荧光光谱研究

白茯苓凝集素（ＳＬＬ）分子中含有４个色氨酸（Ｔｒｐ）残基,ＮＢＳ修饰测得这４个Ｔｒｐ残基位于分子表面。ＳＬＬ在天然状态下荧光发射峰位于３３５ｎｍ处,离子强度和温度对其荧光光谱均无明显的影响。ＮＢＳ修饰后的ＳＬＬ失去凝血活性,相应荧光光谱的强度减弱,荧光发射峰发生蓝移,提示ＳＬＬ的构象发生改变。用ＫＩ·ＣｓＣｌ和丙烯酰胺淬灭剂研究ＳＬＬ分子中Ｔｒｐ残基的微环境,发现丙烯酰胺和ＣｓＣｌ能淬灭分子中１００％和５０％的Ｔｒｐ残基的荧光,而ＫＩ完全不能淬灭ＳＬＬ分子中Ｔｒｐ残基的荧光,因此Ｔｒｐ残基周围存在阴离子区,或者Ｔｒｐ残基处于分子表面的疏水环境中。     

复合探针荧光定量PCR方法的建立

为了对特定基因进行实时检测,根据荧光能量转移(FRET)原理,设计及合成了一种新的FRET复合探针,该探针由一条长的荧光杂交探针和短的淬灭探针构成,其中荧光探针5′端接一荧光素分子,3′端接一延伸阻断分子磷酸,淬灭探针3′端连接一个淬灭分子对甲基红,淬灭探针与荧光探针5′端互补,无模板时,该探针杂交形成复合探针。无荧光产生,当有模板时,荧光探针与模板杂交,荧光不能被淬灭,产生的荧光与模板量成正比。根据复合探针的反应原理,研究了该探针的FRET性质及影响因素包括淬灭探针及扩增片段长度、荧光探针与淬灭探针的合适比例及镁离子浓度。实验结果显示淬灭探针及扩增片段长度对复合探针的作用有明显的影响,本实验采用淬灭探针长21个核苷酸,扩增片段长127bp,荧光探针与淬灭探针的合适比例为1：1,镁离子浓度为3mmo1／L,可获得最佳的反应体系;该复合探针合成简单,淬灭彻底,具有良好的准确性与特异性,敏感性达10^2拷贝,并具有较宽的动力学定量范围,可对10^2—10^9拷贝范围内的待检样品进行准确的定量。复合探针技术可应用于病毒感染水平、转基因拷贝数及单核苷酸多态性等检测。     

硫酸粘菌素对碱性磷酸酶部分性质的影响

硫酸粘菌素对碱性磷酸酶有抑制作用,用Dixon作图求出抑制常数为Ki=4.88×10-6mol/L,抑制类型为非竞争性抑制.通过紫外吸收光谱,发现硫酸粘菌素引起碱性磷酸酶的空间结构发生改变;由荧光光谱变化,发现硫酸粘菌素对碱性磷酸酶有荧光淬灭作用,淬灭机理是因能量转移而引起的静态淬灭.     

An improved surface-based method for DNA computation

DNA computing is a novel method for solving a class of intractable computational problems, in which the computing time can grow exponentially with problem size. Up to now, many accomplishments have been achieved to improve its performance and increase its reliability, among which a surface-based method is an efficient candidate. In this paper, the surface-based approach proposed by Liu, Q., Wang, L., Frutos, A.G., Condon, A.E., Corn, R.M., and Smith, L.M., 2000, DNA computing on surfaces. Nature 403, 175-179 is analyzed and an improved surface-based method for DNA computation (i.e. the hybrid DNA/optical computing method) is proposed. Compared with Liu et al.'s approach, our method has some significant advantages such as low cost, short operating time, reusable surface and simple experimental steps. Moreover, the concept of combining easily patterned DNA computing steps with equally parallel, but generally uniform and not easily patterned optical computing steps is an important new direction.     

Allograft selection for distal femur through cutting contour registration

Allograft reconstruction is an acceptable procedure for the recovery of normal anatomy after the bone tumor resection. During the past few years, several automated methods have been proposed to select the best anatomically matching allograft from the virtual donor bone bank. The surface-based automated method uses the contralateral healthy bone to obtain the normal surface shape of the diseased bone, which could achieve good matching of the defect and the selected allograft. However, the surface-based method focuses on the matching of the whole bone so that the matching of the contact surface between the allograft and the recipient bone may not be optimal. To deal with the above problem, we propose a cutting contour based method for the allograft selection. Cutting contour from the recipient bone could reflect the structural information of the defect and is seldom influenced by tumor. Thus the cutting contour can be used as the matching template to find the optimal alignment of the recipient bone and the allograft. The proposed method is validated using the data of distal femurs where bone transplantation is commonly performed. Experimental results show that the proposed method generally outperforms the surface-based method within modest extra time. Overall, our contour-based method is an effective complementary technique for allograft selection in the virtual bone bank.     

Detection of epileptogenic cortical malformations with surface-based MRI morphometry

Magnetic resonance imaging has revolutionized the detection of structural abnormalities in patients with epilepsy. However, many focal abnormalities remain undetected in routine visual inspection. Here we use an automated, surface-based method for quantifying morphometric features related to epileptogenic cortical malformations to detect abnormal cortical thickness and blurred gray-white matter boundaries. Using MRI morphometry at 3T with surface-based spherical averaging techniques that precisely align anatomical structures between individual brains, we compared single patients with known lesions to a large normal control group to detect clusters of abnormal cortical thickness, gray-white matter contrast, local gyrification, sulcal depth, jacobian distance and curvature. To assess the effects of threshold and smoothing on detection sensitivity and specificity, we systematically varied these parameters with different thresholds and smoothing levels. To test the effectiveness of the technique to detect lesions of epileptogenic character, we compared the detected structural abnormalities to expert-tracings, intracranial EEG, pathology and surgical outcome in a homogeneous patient sample. With optimal parameters and by combining thickness and GWC, the surface-based detection method identified 92% of cortical lesions (sensitivity) with few false positives (96% specificity), successfully discriminating patients from controls 94% of the time. The detected structural abnormalities were related to the seizure onset zones, abnormal histology and positive outcome in all surgical patients. However, the method failed to adequately describe lesion extent in most cases. Automated surface-based MRI morphometry, if used with optimized parameters, may be a valuable additional clinical tool to improve the detection of subtle or previously occult malformations and therefore could improve identification of patients with intractable focal epilepsy who may benefit from surgery.     

DNA computation model to solve 0-1 programming problem

0-1 programming problem is an important problem in opsearch with very widespread applications. In this paper, a new DNA computation model utilizing solution-based and surface-based methods is presented to solve the 0-1 programming problem. This model contains the major benefits of both solution-based and surface-based methods; including vast parallelism, extraordinary information density and ease of operation. The result, verified by biological experimentation, revealed the potential of DNA computation in solving complex programming problem.     

Three-dimensional trabecular alignment model

Trabecular alignment theory has been used to quantify Wolff's Law of bone remodeling. A three-dimensional finite element scheme was developed to analyze the bone remodeling phenomenon. The mathematical model proposed by Mullender et al. and later modified by Smith et al. was adopted to simulate the surface-based trabecular resorption and formation processes. Enhancements incorporated into the previous model include: mapping into three-dimensions, controlling the remodeling signal's passage through marrow, controlling the finite distance the signal may pass through the bone matrix, and including non-bone material in the finite element model. After the model is explained and thoroughly studied, three-dimensional implant surface geometries are simulated.     

High throughput solution-based measurement of antibody-antigen affinity and epitope binning

Advances in human antibody discovery have allowed for the selection of hundreds of high affinity antibodies against many therapeutically relevant targets. This has necessitated the development of reproducible, high throughput analytical techniques to characterize the output from these selections. Among these characterizations, epitopic coverage and affinity are among the most critical properties for lead identification. Biolayer interferometry (BLI) is an attractive technique for epitope binning due to its speed and low antigen consumption. While surface-based methods such as BLI and surface plasmon resonance (SPR) are commonly used for affinity determinations, sensor chemistry and surface related artifacts can limit the accuracy of high affinity measurements. When comparing BLI and solution equilibrium based kinetic exclusion assays, significant differences in measured affinity (10-fold and above) were observed. KinExA direct association (k_a) rate constant measurements suggest that this is mainly caused by inaccurate k_a measurements associated with BLI related surface phenomena. Based on the kinetic exclusion assay principle used for KinExA, we developed a high throughput 96-well plate format assay, using a Meso Scale Discovery (MSD) instrument, to measure solution equilibrium affinity. This improved method combines the accuracy of solution-based methods with the throughput formerly only achievable with surface-based methods.     

Scalability of the surface-based DNA algorithm for 3-SAT

Since Adleman first proposed DNA computing for the Hamiltonian path problem, several authors have reported DNA computing for 3-SAT. Previous research presented DNA computing on surfaces and demonstrated how to solve a four-variable four-clause instance of 3-SAT, and claimed that the surface-based approach was designed to scale up to larger problems. In this paper we establish an error model for the incomplete "mark" and imperfect "destroy" operations. By using the error model we argue that no matter how large the "mark" and "destroy" rates are we can always give satisfiable instances of 3-SAT such that no DNA strands remain on the surface at the end of the computation. By the surface-based approach the satisfiable instances of 3-SAT would be misdetermined to be unsatisfiable. Thus, the error leads to an incorrect result of the SAT computation. Furthermore, given the "mark" rate p and the "not-destroy" rate rho, we find that the approach can only solve at most N-variable instances of 3-SAT problems, where N=[(2+beta(2)+2+2 square root beta (2))/beta(2)] in which beta=1-1/(p+rhoq) and q=1-p and [a] is the greatest integer less than a or equal to a.     

The surface-based approach for DNA computation is unreliable for SAT

Previous research presented DNA computing on surfaces, which applied to each clause three operations:"mark","destroy", and "unmark", and demonstrated how to solve a four-variable four-clause instance of the 3-SAT. It was claimed that only the strands satisfying the problem remained on the surface at the end of the computation and the surface-based approach was capable of scaling up to larger 3-SAT problems. Accordingly, the identities of the strands were only determined in the"readout" step for the correct solutions to the problem without checking if the strands really satisfied the problem. Thus, based on the claim above, the surface-based approach became a polynomial-time algorithm. In this paper, we show that for some instance of SAT, at the end of the computation all the remaining strands falsify the instance. However, by the previous claim all the strands falsifying the problems would be regarded as the correct solutions to the problems. Therefore, the DNA computing on surfaces is unreliable. For this reason, it is necessary to add a "verify" step after the "readout" step to check if the strands remaining on the surface at the end of the computation really satisfy the problem.