催化抗体研究进展

本文评述现代化学和免疫学领域中的一项重大发现——“催化抗体”(Catalytic Antibody,或Abzyme)及其研究进展,回顾了这一发现的分子生物学基础和背景,比较了抗体与酶之间的异同,阐述了赋予抗体催化性质的一般战略,总结了抗体-催化遵循的重要规律,预示了免疫化学的一个新分支学科——Abzymology(译名为“抗体酶学”)的崛起.     

抗体催化的有机化学反应

抗体催化的有机化学反应周建峰（江苏淮阴师专化学系２２３００１）近年来,在生物有机化学领域中出现了一个活跃的研究方向──催化抗体（Ａｂｚｙｍｅ或ＣａｔａｌｖｔｉｃＡｎｔｉｂｏｄｉｅｓ）,它是利用生物学与化学的成果在分子水平上交叉渗透研究的产物。由于催化...     

催化抗体

动物的免疫系统根据过渡态相似物的结构,量体裁衣式地产生形状、电性、酸碱性互补的催化抗体,它能高选择性高效率地催化化学反应。采用单克隆、多克隆、基因工程等技术,众多的性能优越的催化抗体相继问世,它标志着催化剂的研究进入了一个崭新的领域,表明模拟酶的研究发生了质的飞跃,预示着极其广阔的应用前景。     

催化抗体化学的新进展

近年来，生物有机化学中的最新成就之一是催化抗体的开发成功。其蓬勃发展的势头，方兴未艾。本文介绍催化抗体的多样性及其根源、产生方法、催化特性及发展前景。     

催化抗体在有机化学中的应用

介绍了催化抗体在有机化学中的应用，重点阐述了催化抗体的高度特异性。     

催化抗体的催化效率优化和展望

综述了改进催化抗体催化效率的可能方法 ,包括较好的半抗原、广泛的筛选、工程化及选择优化等     

具有谷胱甘肽过氧化物酶活性的催化抗体的研究：Ⅰ.谷胱甘肽特征全抗…

用２,４－二硝基氯苯保护谷胱甘肽的巯基制出半抗原,再通过戊二醛共价反应使其与牛血清白蛋白表面的氨基结合,经超胶ＡｃＡ５４凝胶层析纯化,制备出有较强免疫原性的谷胱甘肽全抗原,用元素分析、红外光谱及核磁共振表征了半抗原的结构,电泳分析得全抗原分子量平均为８７０００道尔顿,光谱分析及圆二色谱研究表明其有较强的可强,紫外及荥光特征吸收,且抗原结构的紧密性增强。     

《分子催化》简介

《分子催化》是由中国科学院兰州化学物理研究所主办、中国科学院主管、科学出版社出版的向国内外公开发行的学术性刊物。主要报道有关分子催化方面最新进展与研究成果。辟有学术论文、研究简报、研究快报及综合述评等栏目。内容侧重于配位催化、酶催化、光助催化、催化过程中的立体化学问题、催化反应机理与动力学、催化剂表面态的研究及量子化学在催化学科中的应用等。工业催化过程中的均相催化     

具有谷胱甘肽过氧化物酶活性的催化抗体的研究Ⅰ．谷胱甘肽特征全抗原的合成、表征及性质

用２，４－二硝基氯苯保护谷胱甘肽的巯基制出半抗原，再通过戊二醛共价反应使其与牛血清白蛋白表面的氨基结合，经超胶ＡｃＡ５４凝胶层析纯化，制备出有较强免疫原性的谷胱甘肽全抗原。用元素分析、红外光谱及核磁共振表征了半抗原的结构。电泳分析得全抗原分子量平均为８７０００道尔顿。光谱分析及圆二色谱研究表明其有较强的可见、紫外及荧光特征吸收，且抗原结构的紧密性增强。     

A catalytic antibody against a tocopherol cyclase inhibitor

The cyclic ammonium cation 5 and its guanidinium analogue 4 are inhibitors of tocopherol cyclase. Monoclonal antibodies were raised against protein conjugates of the haptens 1-3 and screened for catalytic reactions with alkene 8, a short chain analogue of the natural substrate phytyl-hydroquinone 6, and its enol ether analogues 10a,b. Antibody 16E7 raised against hapten 3 was found to catalyze the hydrolysis of Z enol ether 10a to form hemiacetal 12 with an apparent rate acceleration of k(cat)/k(uncat)=1400. Antibody 16E7 also catalyzed the elimination of Kemp's benzisoxazole 59. The absence of cyclization in the reaction of enol ether 10a was attributed to the competition of water molecules for the oxocarbonium cation intermediate within the antibody binding pocket. Hapten and reaction design features contributing to this outcome are discussed. Antibody 16E7 provides the first example of a carboxyl group acting both as an acid in an intrinsically acid-catalyzed process and as a base in an intrinsically base-catalyzed process, as expected from first principles. In contrast to the many examples of general-acid-catalyzed processes known to be catalyzed by catalytic antibodies, the specific-acid-catalyzed cyclization of phytyl-hydroquinone 6 or its analogue 8 still eludes antibody catalysis.     

Immune recognition,antigen design,and catalytic antibody production

Catalytic antibodies have been developed by experimental approaches exploiting the analogy between antibody-antigen and enzyme-substrate interaction. Haptens have been prepared to model the electrostatic or geometric attributes of a reaction’s transition state and to induce combining sites having appropriate catalytic residues. The relative merits of these design strategies may be gleaned from the apparent activities and efficiencies of the respective catalysts. The implications of screening strategies on the kinetic characteristics of the resulting abzymes are also considered. Combining-site hypermutation provides the variation in the antibody repertoire from which high-affinity clones are selected. The same mechanism can also lead to a subset of antibodies with reduced hapten affinity, but improved catalytic activity. This possibility has not been adequately characterized, but is suggested by a number of considerations. These include the unexplained efficiency and diversity of mechanisms utilized by various antibody catalysts, and the observed catalytic activity of antibodies found in autoimmune serum. This article attempts to assess critically the evidence for rational design of catalytic activity in antibodies. Correlations among abzymes and their relevant models could lead to revised or novel strategies for producing better catalysts.     

钴卟啉过氧中间体向环己烷夺氢的密度泛函计算

采用密度泛函理论计算方法模拟了简单钴卟啉过氧中间体PCo-O2与环己烷C6H12的作用,分析了反应路径中各驻点能量和反应过渡态分子构型。研究结果表明,PCo-O2向底物环己烷夺氢的反应可以延正方向进行,二线态PCo-O2更具反应活性,反应过程中Co-O键得到加强,O-O键被削弱。依据理论计算结果,探讨了四苯基钴卟啉催化环己烷氧化生成环己醇和环己酮的反应机理,指出反应延Lyons高价金属氧代物机理生成环己醇,而反应循环中产生的烷基自由基可以延烷基过氧化过渡金属配合物反应机理进行生成环己酮。     

Comparison of antibody and albumin catalyzed hydrolysis of steroidalp-nitrophenylcarbonates

A monoclonal antibody (MAb) was produced against thep-nitrophenylphosphate derivative of 3α,5β-lithocholic acid, a transition-state analog for hydrolysis of a steroidalp-nitrophenylcarbonate. The indicated reaction was catalyzed by this Ab with kinetic constants k_cat = 4.0 × 10^-2min and K_m = 3.3 μM at pH 9.0 and 35°C. The Ab also hydrolyzed the isomericp-nitrophenylcarbonate of 3β,5β-lithocholic acid with k_cat = 8.4 × 10^-2/min and K_m = 1.0 μM. Bovine serum albumin (BSA) was found to catalyze the same reactions with similar turnover rates and Michaelis constants of 15 and 14 μM, respectively. Although the BSA-catalyzed reaction was only weakly inhibited by the phosphate ester TSA (IC₅₀ ca. 40 μM), the Ab-catalyzed reaction was completely inhibited at less than 1 μM of the TSA. The relative rates and efficiencies of the MAbcatalyzed and BSA-catalyzed reactions are discussed in the context of the hydrophobic sites and intrinsic reactivity of the protein surfaces, and the induction of groups on the Ab to enhance the enzymatic function.     

催化2-(对甲氧苯氧基)-四氢吡喃缩醛键水解的抗体研究

以2-[对-(3-羧基)-丙基氧苯]-亚氨基哌啶(2)为半抗原, 联接载体蛋白后免疫动物, 并经杂交瘤技术获得十株单克隆抗体, 其中一株单抗1D1在pH=5.6, 37℃条件下能催化2-(对-甲氧苯氧基)-四氢吡喃(1)缩醛键水解, 加速37倍。     

Catalytic activity of anti-ground state antibodies,antibody subunits,and human autoantibodies

Catalytic antibodies may be produced over the natural course of antibody-affinity maturation by placement of chemically reactive residues in antibody-active sites by somatic hypermutation or V-D-J-gene rearrangement. This hypothesis has received support from recent observations on the chemical reactivity of antibodies to vasoactive intestinal peptide (VIP), DNA, and steroid-and dinitrophenyl-esters. Recent studies reveal that monoclonal antibodies raised against the ground state of VIP can accelerate the cleavage of peptide bonds. The light-chain (L-chain) subunit of human autoantibodies display increased hydrolytic rate and diminished VIP-binding affinity compared to the parent antibody, consistent with increased turnover owing to weaker binding of the substrate ground state. These observations reveal an essential limitation of catalytic antibodies, i.e., large turnover rates may be associated with diminished substrate specificity. The hydrolysis of VIP by IgG purified by affinity chromatography from asthma patients and nonasthmatic controls was compared. IgG from the majority of asthma patients displayed VIP-hydrolyzing activity. V_max values for IgG from asthmatics tended to be higher than those from the nonasthmatic group. In principle, catalysis by antibodies may be an important mediator of immunological defense, regulation, and autoimmune dysfunction. The verification of these possibilities will require studies that utilize efficient assays of antibody catalysis during experimental immunization and autoimmune disease, as well as mechanistic investigation of catalysis by antibodies and their subunits.     

Binding of active site directed ligands to phospholipase A2: Implications on the molecular constraints and catalytic mechanism

Molecular constraints for the localization of active site directed ligands (competitive inhibitors and substrates) in the active site of phospholipase A2 (PLA2) are characterized. Structure activity relationships with known inhibitors suggest that the head group interactions dominate the selectivity as well as a substantial part of the affinity. Theab initio fitting of the amide ligands in the active site was carried out to characterize the head group interactions. Based on a systematic coordinate space search, formamide is docked with known experimental constraints such as coordination of the carbonyl group to Ca²⁺ and hydrogen bond between amide nitrogen and ND1 of His48. An optimal position for a bound water molecule is identified and its significance for the catalytic mechanism is postulated. Unlike the traditional “pseudo-triad” mechanism, the “Ca-coordinated-oxyanion” mechanism proposed here invokes activation of the catalytic water to form the oxyanion in the coordination sphere of calcium. As it attacks the carbonyl carbon of the ester, a near-tetrahedral intermediate is formed. As the second proton of the catalytic water is abstracted by the ester oxygen, its reorientation and simultaneous cleavage form hydrogen bond with ND1 of His48. In this mechanism of esterolysis, a catalytic role for the water co-ordinated to Ca²⁺ is recognised.     

On the possible reaction pathway for the acylation of AChE-catalyzed hydrolysis of ACh: Semiempirical quantum chemical study

The acylation process in the acetylcholinesterase (AChE)-catalyzed hydrolysis of the neurotransmitter, acetylcholine (ACh), has been determined with the semiempirical quantum chemical calculation method AM1 using the model molecules extracted from the X-ray crystal structure of Torpedo Californica AChE. For the sake of identifying the microfeatures of the mechanism of this reaction, two types of possible mechanisms, stepwise mechanism and cooperative mechanisms, were proposed and studied with AM1 methods. All the model molecules for the possible reactants, intermediates, transition states, and products in the reaction pathways of the two mechanisms were obtained. Energy profiles, the structural properties of the transition states, indicate that the acylation of AChE-catalyzed hydrolysis of ACh adopts the cooperative mechanism, i.e., the proton transfer from Ser220 of AChE to His440 occurs simultaneously with the nucleophilic attack of Ser200 to the carbonyl carbon atom of ACh. This result is in agreement with the kinetic data and the secondary isotope effects of AChE-catalyzed reactions. © 1998 John Wiley & Sons, Inc. Int J Quant Chem 70: 515–525, 1998     

X-Ray structures of two families of hydrolytic antibodies

The catalytic mechanisms of two esterase-like catalytic antibodies (Abs) have been determined, based on kinetic data and on structures of the complexes with transition-state analogs and with a stable substrate analog of the reactions they catalyze. Both Abs stabilize the oxyanion intermediate close to the transition state in ester hydrolysis. The different geometries of the hydrogen bonds that participate in this stabilization account for most of the difference between the efficiencies of these two Abs.