Biochemistry And Pharmacology of Fluorinated Imidazoles

氟化咪唑的生物化学和药理学

• 批准号: 7734050
• 负责人: KENNETH L KIRK
• 金额: $ 41.39万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7734050
• 关键词: Amides; Anabolism; Anthrax disease; Antigens; Bacillus anthracis; Binding; Biochemical; Biochemistry and Pharmacology Cancer Activity; Biological; Biological Process; Carnosine; Catalytic Domain; Cell Death; Complex; Data; Dependence; Dipeptides; Dissociation; Enzymes; Evaluation; Event; Fluorine; Future; Histamine; Histidine; Hydrogen; Imidazole; Immune response; In Vitro; Label; Lead; Light; Measurement; Mediating; Mediator of activation protein; Monitor; Motion; Nerve; Neurotransmitters; Object Attachment; Pathogenesis; Play; Procedures; Process; Property; Proteins; Purines; Range; Reporting; Research; Role; Signal Transduction; Structure; System; Titrations; Toxic effect; Work; analog; anthrax lethal factor; anthrax protective factor; anthrax toxin receptors; beta-Alanine; blood glucose regulation; design; hydroxyl group; in vivo; interest; protein structure; protonation; purine; receptor; research study; riboside; size; tool

In light of the many important functions of the imidazole ring, it is not surprising that many of the ring-fluorinated analogues proved to have quite interesting biological properties. 2-Fluorohistidine is a notable example, showing a range of activities that may or may not be related to the ability of this analogue to be incorporated into protein in vivo. We recently have re-initiated efforts to define the mechanisms by which this analogue exerts its effects. Fluorohistidine: Biochemical Incorporation of Fluorohistidine into Proteins. Work is continuing in a collaborative project on incorporation of 2-fluoro-L-hisitidine into anthrax protein as a tool to study details of the mechanism of toxicity. The pathogenesis of Bacillus anthracis relies in part upon a pH dependent conversion of the anthrax protective antigen (PA) from a heptameric prepore to a pore. Lowering the pH in vitro from 8 to 7 can induce pore formation by protective antigen. However, a pH of 6 is required in vivo in the presence of the receptor to induce pore formation and dissociation from the receptor. The pH dependence of pore formation in the presence of the anthrax ANTXR2 receptor is consistent for the titration of His residues. There are several His residues in the domain of PA63 critical for pore formation as well as a His residue on CMG2. Thus, protonation of His in ANTXR2 andor PA and subsequent conformational changes have been suggested as critical events in pore formation. To investigate this,2FHis has been incorporated into both the receptor (ANTXR2) and PA83. Since the imidazole pKa of 2FHis is about 1, protonation of 2FHis will not occur under the pH changes that lead to pore formation. This will allow determination of the importance of His protonation in anthrax infectivity. As reported previously, the presence of 2FHis in the anthrax receptor ANTXR2 had no effect on pore formation mediated by native PA. Thus, the proposal that a CMG2-His protonation repels an Arg in PA63 causing conformational changes that lead to pore formation appears unlikely. Since 2FHis-ANTXR2 possesses only a single resonance in the 19F NMR spectrum (there is only one histidine in ANTXR2), this signal allowed us to monitor association-dissociation to PA as a function of pH using 19F-NMR. In these experiments, 2FHis-ANTXR2 is added to (PA63)7, resulting in a significant decrease in the intensity of the 2-FHis resonance, presumably from the slower tumbling motion of the complex. As the pH is lowered to 5, an increase in the intensity of the fluorine resonance occurs, indicating that ANTXR2 dissociates from PA upon formation of the transmembrane pore. These data are consistent with of others, and provide the first direct evidence that dissociation of the receptor occurs upon pore formation. In addition, the pH for pre-pore to pore conversion was not altered for 2FHis-labelled PA in the absence of receptor. However, pore formation by 2FHis-labelled PA from heptameric pre-pore (2-FHis PA63)7, in the presence of the receptor was blocked. In addition, translocation experiments show that pores formed from (2-FHis PA63)7, in the absence of receptor are unable to translocate LF. In addition, 2FHis PA is unable to mediate cell death in vivo. From these results it seems likely that protonation of residues in PA causes conformational changes that lead to pore formation and that binding to the receptor inhibits these changes. The mechanism by which ANTXR2 blocks pore formation mediated by 2FHis PA is not obvious. One possibility is that 2FHis PA forms a more stable complex with the receptor, thus blocking the dissociation that accompanies heptamer formation and pore formation. It also is not clear why pores that are formed in the absence of receptor protein are unable to translocate lethal factor. Work in progress is designed to identify the critical His residues in these processes. NMR measurements using 2-FHis121-ANTXR2 have recently provided direct evidence that protonation events in the protective antigen prepore result in dissociation from the anthrax toxin receptor at acidic pH. Synthesis of 2-FHis and 4-FHis containing analogs of carnosine The dipeptide carnosine (beta-alanyl-His) has many important biological functions, including a possible role in the regulation of blood glucose through control of autnonomic nerves. We have embarked on a project to prepare carnosine derivatives wherein histidine is replaced with 2-FHis or 4-FHis. Normal amide bond forming procedures were used to to prepare the dipeptides from beta-alanine and 2- and 4-FHis. Future plans include evaluation of these new analogues in carnosine utilizing systems.

鉴于咪唑环的许多重要功能，许多环氟化类似物被证明具有相当有趣的生物学特性就不足为奇了。2-氟组氨酸是一个值得注意的例子，它显示了一系列的活性，这些活性可能与这种类似物在体内并入蛋白质的能力有关，也可能与之无关。我们最近重新开始努力，以确定这种类似物发挥作用的机制。 氟组氨酸： 氟组氨酸在蛋白质中的生物化学掺入 将2-氟-L-组氨酸掺入炭疽蛋白作为研究毒性机制细节的工具的合作项目仍在继续。炭疽杆菌的致病机制部分依赖于炭疽保护性抗原(PA)从七聚体前孔到孔洞的pH依赖转换。在体外将pH从8降低到7可以通过保护性抗原诱导孔洞形成。然而，在体内，在受体存在的情况下，需要pH为6才能诱导孔洞形成和从受体解离。在炭疽ANTXR2受体存在下，孔道形成的pH依赖性与His残基的滴定是一致的。在PA63区域有几个His残基对孔隙形成至关重要，在CMG2上也有一个His残基。因此，His在ANTXR2和或PA中的质子化和随后的构象变化被认为是孔隙形成的关键事件。为了研究这一点，2FHis已经被整合到受体(ANTXR2)和PA83中。由于2FHis的咪唑pKa约为1，2FHis在pH变化下不会发生质子化而导致孔道形成。这将有助于确定他的质子化在炭疽传染性中的重要性。 正如先前报道的那样，炭疽受体ANTXR2中2FHis的存在对天然PA介导的孔道形成没有影响。因此，认为CMG2-His质子化排斥PA63中的Arg导致构象变化从而导致气孔形成的说法似乎不太可能。由于2FHis-ANTXR2在19F核磁共振谱中只有一个共振(ANTXR2中只有一个组氨酸)，这一信号使我们能够使用19F-核磁共振监测与PA的缔合-解离作为pH的函数。在这些实验中，2FHis-ANTXR2被添加到(PA63)7中，导致2-FHis共振的强度显著降低，推测是由于络合物较慢的翻滚运动。当pH降低到5时，氟共振的强度增加，表明ANTXR2在形成跨膜孔时与PA解离。这些数据与其他数据是一致的，并提供了第一个直接证据，表明该受体在孔隙形成时发生解离。 此外，在没有受体的情况下，2FHis标记PA的预孔向孔转化的pH没有改变。然而，在受体存在的情况下，2FHis标记的PA从七聚体前孔(2-FHis PA63)7形成的孔道被阻断。此外，易位实验表明，在没有受体的情况下，(2-fHis PA63)7形成的孔道不能易位LF。此外，2FHis PA在体内不能介导细胞死亡。从这些结果来看，PA中残基的质子化似乎可能导致构象变化，从而导致孔形成，而与受体的结合抑制了这些变化。ANTXR2阻断2FHis PA介导的成孔作用机制不明显。一种可能性是2FHis PA与受体形成更稳定的复合体，从而阻止伴随七聚体形成和孔形成的解离。同样不清楚的是，为什么在缺乏受体蛋白的情况下形成的毛孔无法转移致死因子。正在进行的工作旨在确定这些过程中关键的组氨酸残留物。 使用2-FHis121-ANTXR2的核磁共振测量最近提供了直接证据，表明保护性抗原前孔中的质子化事件导致在酸性pH下从炭疽毒素受体解离。 含肌肽类似物的2-FHIS和4-FHIS的合成 二肽肌肽(β-丙氨酰-组氨酸)具有许多重要的生物学功能，包括可能通过控制自体神经来调节血糖。我们已经开始了一个项目来制备肌肽衍生物，其中组氨酸被2-FHIS或4-FHIS取代。以β-丙氨酸、2-FHIS和4-FHIS为原料，采用常规的酰胺键形成方法制备二肽类化合物。未来的计划包括在肌肽利用系统中评估这些新的类似物。

项目成果

Behavior of fluorinated analogs of L-(3,4-dihydroxyphenyl)alanine and L-threo-(3,4-dihydroxyphenyl)serine as substrates for Dopa decarboxylase.

L-(3,4-二羟基苯基)丙氨酸和L-苏-(3,4-二羟基苯基)丝氨酸的氟化类似物作为多巴脱羧酶底物的行为。

• DOI: 10.1016/s0006-291x(02)00643-5
• 发表时间: 2002
• 期刊: Biochemical and biophysical research communications
• 影响因子: 3.1
• 作者: BorriVoltattorni,Carla;Bertoldi,Mariarita;Bianconi,Silvia;Deng,Wei-ping;Wong,Kelli;Kim,InHo;Herbert,Brian;Kirk,KennethL
• 通讯作者: Kirk,KennethL

An Enantioselective Synthesis of (S)-4-Fluorohistidine.

(S)-4-氟组氨酸的对映选择性合成。

• DOI: 10.1016/j.jfluchem.2008.04.011
• 发表时间: 2008
• 期刊: Journal of fluorine chemistry
• 影响因子: 1.9
• 作者: Hajduch,Jan;Cramer,JohnC;Kirk,KennethL
• 通讯作者: Kirk,KennethL

Synthesis of (E)- and (Z)-alpha,beta-Difluorourocanic Acid.

(E)-和(Z)-α,β-二氟尿刊酸的合成。

• DOI: 10.1016/j.jfluchem.2007.09.006
• 发表时间: 2008
• 期刊: Journal of fluorine chemistry
• 影响因子: 1.9
• 作者: Hajduch,Jan;Dolenský,Bohumil;Yoshida,Shinichi;Fan,Junfa;Kirk,KennethL
• 通讯作者: Kirk,KennethL

Gc protein (vitamin D-binding protein): Gc genotyping and GcMAF precursor activity.

• 发表时间: 2005-11
• 期刊: Anticancer research
• 影响因子: 2
• 作者: H. Nagasawa;Y. Uto;Hideyuki Sasaki;Natsuko Okamura;Aya Murakami;S. Kubo;K. Kirk;H. Hori

Simple synthesis of alpha-cyano-alpha-fluoro-p-tolylacetic acid (CFTA), a new efficient chiral derivatizing agent.

新型高效手性衍生剂α-氰基-α-氟-对甲苯乙酸（CFTA）的简单合成。

• 发表时间: 1999
• 期刊: Enantiomer
• 作者: Takeuchi,Y;Konishi,M;Hori,H;Takahashi,T;Kirk,KL
• 通讯作者: Kirk,KL