SYNTHESIS AND PROPERTIES OF ORGANOCOBALT COMPLEXES

关于B12依赖的重要未解问题 酶的过程是：构象变化是什么 它们促进了Co-C键的均质裂解速率 辅酶B12增加了10-10倍，有利于均质 B12全酶中的过度异源裂解？为什么Co- N(DMBz)键这么长，DMBz在其中扮演什么角色 流程？如果酶对重排的贡献 工艺(如甲基丙二酰辅酶A到琥珀酰辅酶A)受到限制 为了确保长寿命底物自由基的存在，如何做 这些酶阻止了与B12？这些和 由于几个原因，其他问题还没有得到完全回答。 依赖于B12的酶很大，可以产生X射线和一些 光谱研究很困难。辅因通常紧密地存在于 绑在探头无法接触到的受保护的酶口袋中 试剂。只有四种烷基钴钴的X射线结构包括 辅酶B12和甲基B12已有报道。这些分子 是复杂和独特的，光谱方法受到以下限制 这种复杂性和缺乏背景数据。因此， 研究很好的辅酶类似物的构象是未知的。 这些有限信息的一个后果是，几乎没有 明确结构和光谱之间的关系。还有，还有 仍然是一个不充分的背景，用来评估立体 导致Co-C键断裂的因素。以继续解决 针对这些问题，我们提出了一个三管齐下的方法。详细 (I)烷基钴化合物的研究；(Ii)合成有机钴的研究 以及(Iii)核糖核苷酸还原酶(RR)。 烷基钴胺类化合物的可能构象范围为 通过合成改性的或立体应变的 烷基钴化合物(钴胺、钴酰胺和其他侧链-- 修饰的衍生物)和附加的有机钴化合物 空间或电子修饰的赤道配体 科萨洛夫和“科斯塔”系统。结构的影响 在这些系统中引起的变化将通过X射线进行评估 结晶学，现代光谱方法(CD、2D和 HMQC核磁共振、拉曼)以及反应速率和机理 研究(DMBz解离、配体交换)。应用程序 其中一些方法已经产生了丰富的信息和 在此赠款期间提供了新的见解。这些方法将 可推广到RR的研究。RR相对较小，具有 松散绑定的可访问、B12站点和作用机制 与铁酶有关。因此，RR具有特殊的优势和 意义。钴和钴在烷基键合中的空间位阻 辅酶类似物中的腺嘌呤也将发生变化， 类似物和辅酶B12与RR的相互作用将是 调查过了。对于核磁共振研究，我们将使用钴胺类药物 含有同位素浓缩点的。光谱(Cd，Vis， 核磁共振(2D、HMQC))，背景待开发一般 可用于其他B12全酶和B12转运的研究 系统。

Important unanswered questions concerning B12-dependent enzymic processes are: What are the conformational changes which promote the rate of homolytic cleavage of the Co-C bond of coenzyme B12 by a factor of 10-10 and which favor homolytic over heterolytic cleavage in B12 holoenzymes? Why is the Co- N(DMBz) bond so long and what is the role of DMBz in these processes? If the contribution of the enzyme to rearrangement processes (e.g. methylmalonyl-CoA to succinylCoA) is restricted to insuring the existence of long-lived substrate radicals, how do the enzymes prevent radical combination with B12? These and other questions have not been fully answered for several reasons. The B12-dependent enzymes are large, making X-ray and some spectroscopic studies difficult. The cofactor generally lies tightly bound in a protected enzyme pocket inaccessible to probe reagents. Only four alkylcobaltcorrin X-ray structures including coenzyme B12 and methyl B12 have been reported. The molecules are complex and unique and spectroscopic methods are limited by this complexity and by a lack of background data. Therefore, the conformations of well studied coenzyme analogues are unknown. One consequence of this limited information is that there are few clear relationships between structure and spectra. Also, there remains an inadequate background with which to assess the steric factors leading to Co-C bond cleavage. To continue to address these questions, we propose a three-pronged approach. Detailed studies of (i) alkylcobaltcorrins, (ii) synthetic organocobalt species, and (iii) ribonucleotide reductase (RR) are contemplated. The range of conformations possible for alkylcobalamins will be defined via the synthesis of modified or sterically strained alkylcobaltcorrins (cobalamins, cobinamides and other side chain- modified derivatives) and additional organocobalt compounds with sterically or electronically modified equatorial ligands based on the Cosaloph and "Costa" systems. The effect of structural changes induced in these systems will be assessed by X-ray crystallography, modern spectroscopic methods (CD, 2D and HMQC NMR, Raman) and by reaction rates and mechanistic studies (DMBz dissociation, ligand exchange). Application of some of these methods has produced a wealth of information and provided new insight during this grant period. These methods will be extended to the study of RR. RR is relatively small, with a loosely bound accessible, B12 site and a mechanism of action related to the Fe enzymes. Thus, RR has special advantages and significance. Steric strain in the alkyl linkage between Co and adenine in coenzyme analogues will also be varied and the interaction of the analogues and coenzyme B12 with RR will be investigated. For NMR studies, we will employ cobalamins containing isotopically enriched sites. The spectroscopic (CD, vis, NMR (2D, HMQC)), background to be developed will be generally useful in studies on other B12 holoenzymes and on B12 transport systems.