HANDLING OF THIYL METALLOENZYME CENTERS FOR 95 & 140 GHZ EPR STUDIES

95 的硫酰金属酶中心的处理

• 批准号: 6120622
• 负责人: JOANNE STUBBE
• 金额: $ 0.27万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-04-15 至 1999-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6120622
• 关键词: biological products; biomedical equipment development; biomedical resource; enzymes; technology /technique

The ribonucleoside triphosphate reductase (RTPR) from Lactobacillus leichmannii requires adenosylcobalamin (AdoCbl) as a cofactor to catalyze the conversion of nucleotides to deoxynucleotides. RTPR has previously been shown to catalyze the homolytic cleavage of the carbon-cobalt bond of AdoCbl, and the resulting paramagnetic species has been characterized by rapid freeze-quench X-band and Q-band spectroscopy (Orme-Johnson, W. Fl.; Beinert, H.; Blakeley, R.L., J. Biol. Chem. 1974, 249, 2338-2343. Licht, S.; Gerfen, G.J.; Stubbe, J. Science 1996, 271, 477-481). Simulations indicate a two-spin-site system with both cobalt ion and thiyl radical (Gerfen, G.J.; Licht, S.; Willems, J.P.; Hoffman, B.R.; Stubbe, J. JACS 1996, 18:2338-2343), but higher EPR frequencies are needed for definitive conclusions on this and other B12-dependent enzyme systems. Sample preparation, handling, and loading is challenging for the small sample tubes (less than 1 mm ID) of the 95 GHz and 140 GHz EPR spectromete rs at IERC and MIT, respectively. This sample handling problem is being addressed.

核糖核苷三磷酸还原酶（RTPR）来自 莱氏乳杆菌需要腺苷钴胺素（Adenosylcobalamin，ACBl）作为 辅因子催化核苷酸转化为 脱氧核苷酸 RTPR先前已被证明可以催化 均裂的碳-钴键的C_2Cbl，和 所得到的顺磁性物质的特征在于快速的 冷冻淬火X-带和Q-带光谱（Orme-Johnson，W.佛罗里达州; Beinert，H.; Blaucet，R.L.，J. Biol. Chem. 1974，249，2338-2343。 Licht，S.; Gerfen，G.J.; Stubbe，J. Science 1996，271，477-481）。 模拟表明，一个双自旋位系统与钴离子和 硫基自由基（Gerfen，G. J.; Licht，S.; Willems，J.P.;霍夫曼，B.R.; Stubbe，J. JACS 1996，18：2338-2343），但是更高的EPR频率是 需要对这种和其他B12依赖性 酶系统 样品制备、处理和装载是 对于95个样品管中的小样品管（小于1 mm ID）具有挑战性 140 GHz和140 GHz EPR谱仪，分别在IERC和MIT。 正在解决该样品处理问题。