生物化学资料：chapter 9 Tricarboxylic Acid Cycle

1、Biochemistry,Chapter 9 Tricarboxylic Acid Cycle,The decarboxylation of pyruvate, and the oxidation of acetate takes place in the mitochondrial matrix,ATP synthesis takes place in the inner membrane,Glucose Pyruvate Acetal-CoA TCA Cycle,NAD,NADH +H,CoA-SH,CO2,CO2 + H2O,Glycolysis,Yielding of acetyl-C

2、oA,acetyl-CoA is oxidized,TCA,The Nobel Prize in Physiology or Medicine 1953,for his discovery of the citric acid cycle,尿素循环,Pyruvate is transported into mitochondria（线粒体） then oxidized to acetyl-CoA (releasing CO2) before entering the citric acid cycle,Pyruvate is oxidized to acetyl-CoA,丙酮酸脱氢酶系,thr

3、ee enzymes,six coenzymes,PDH complex,pyruvate dehydrogenase (E1,dihydrolipoyl transacetylase (E2,dihydrolipoyl dehydrogenase (E3,CoA-SH NAD+/NADHH+ Lipoate (硫辛酸） FAD TPP Mg2,Regular proteins,Protein kinase Phosphate protein phosphatase,Pyruvate dehydrogenase complex,二氢硫辛酰转乙酰基酶,二氢硫辛酸脱氢酶,硫胺素焦磷酸Vb1,脚气病

4、,The process of oxidation from Pyruvate to acetyl-CoA,丙酮酸与TPP结合并脱羧形成羟乙基TPP。 2. 羟乙基TPP氧化转变成乙酰基同时转移到E2的辅基硫辛酰胺上。 3. 在E2上的乙酰基在E2催化下转移到CoASH 上形成游离的乙酰CoA.从而形成了一个高能硫酯键,4. 还原型的E2将二个SH基H转移到E3的辅酶FAD上形成还原型FADH,5. E3上的还原型的FADH将H交给NAD+形成NADH，E3辅基又形成氧化型的FAD,20,5-6,60,Pyruvate dehydrogenase complex,二氢硫辛酰转乙酰基酶,二氢硫辛

5、酸脱氢酶,硫辛酸,共价 相连,Electron Micrograph of pyruvate dehydrogenase complexes,受蛋白激酶和磷酸蛋白磷酸酶控制磷酸化失活 E1的磷酸化和去磷酸化是使PDH复合体失活和激活的重要方式. 受激素调节 受能荷效应物调节 （变构调节,激活剂,ADP, AMP,E1,CoASH, NAD,E2, E3,抑制剂,ATP, GTP,E1,Acetal-CoA NADHH,E2, E3,Regulation of pyruvate dehydrogenase complex,Regulation of pyruvate dehydrogenase

6、complex,砷化物可与丙酮酸脱氢酶复合物中E2辅基硫辛酰胺的巯基发生共价结合，使还原型硫辛酰胺形成失去催化能力的砷化物,拿破仑之死,砒霜是如何使拿破仑中毒并死亡的呢？Do you know,生活中砒霜的形成,法国人：继承粉末,虾体内所含砷的化学价是五价，一般情况下，五价砷对人体是没有害处的。理论上讲，高剂量的维生素C（一次性摄入维生素C超过500毫克）和五价砷经过复杂的化学反应，会转变为有毒的三价砷（即我们常说的“砒霜”），当三价砷达到一定剂量时可导致人体中毒,维生素C和虾,乙酰CoA,柠檬酸,顺乌头酸,异柠檬酸,a-酮戊二酸,琥珀酰CoA,琥珀酸,苹果酸,延胡索酸,草酰乙酸,Tricar

7、boxylic Acid Cycle,Each acetyl-CoA is completely oxidized to two CO2, generating three NADH, one FADH2, and one GTP; The metabolites contain six, five or four carbons in the cycle,NAD+ and FAD takes the electrons during acetyl-CoA oxidation,The cycle begins with the condensation of acetyle-CoA and o

8、xaloacetate to form citrate,草酰乙酸,柠檬酸,柠檬酸合酶,The hydrolysis of the high energy thioester(硫酯) drives the reaction,催化TCA的第一步反应，反应先生成柠檬酰CoA，再水解为柠檬酸，是放能反应，不可逆；是TCA的第一个调节酶，活性受ATP、NADH、Succinyl CoA(琥珀酰CoA)及长链脂酰CoA的抑制，对于TCA是一个rate-limiting step。 氟乙酰CoA在酶的作用下可与草酰乙酸生成氟柠檬酸，顺乌头酸酶只识别柠檬酸，对氟柠檬酸没有作用，致使TCA中断，这种合成为致死合

9、成(lethal synthesis)。在代谢研究的应用上，曾被广泛用于杀虫剂或灭鼠药的生产 (现已被明令禁止),Citric acid synthase,G0 = 8.4 kJ/mol,G0 = -2.1 kJ/mol,90,4,6,Fluoride citrate（氟柠檬酸） is the special inhbitor of aconitase,Citrate isomerizes to isocitrate (异柠檬酸) via cis-aconitate (顺乌头酸,Aconitase (乌头酸酶,反应具有严格的空间特异性,Reaction 4 and 5,Isocitrate u

10、ndergoes oxidative decarboxylation (氧化脱羧) to form a-ketoglutarate (a-酮戊二酸,Isocitrate is oxidized (第一次脱氢) and decarboxylated.(第一次脱羧) 既有以NAD+为辅酶的异柠檬酸脱氢酶，也有以NADP+为辅酶的异柠檬酸脱氢酶 Positive effectors: ADP, AMP, isocitrate Negative effectors: ATP, NADH,异柠檬酸脱氢酶,Catalyzed by -ketoglutarate dehydrogenase complex;

11、 almost the same as pyruvate dehydrogenase complex; E3 is identical (相同的), E1 and E2 are very similar. 第二次脱氢,第二次脱羧.E1酶不受磷酸共价调节. Reaction requires TPP and lipoate etc(6种辅酶,ketoglutarate undergoes oxidative decarboyxlation to form succinyl-CoA (琥珀酰CoA,a-酮戊二酸脱氢酶系,琥珀酰CoA合成酶 （也称琥珀酸硫激酶,Cleavage of the thi

12、oester bond (硫酯键) in succinyl-CoA is coupled to GTP (or ATP) formation,The third example of substrate-level phosphorylation,琥珀酰CoA,琥珀酸,琥珀酸是对称分子,没有了手性,Cleavage of the thioester bond (硫酯键) in succinyl-CoA is coupled to GTP (or ATP) formation,琥珀酰CoA合成酶,琥珀酸,延胡索酸,Succinate is oxidized to fumarate (延胡索酸,琥

13、珀酸脱氢酶,The free energy change of this reaction is insufficient to reduce NAD+. Succinate Dehydrogenase is directly linked with the electron transport chain,三羧酸循环中唯一与内膜结合的酶，是脱氢酶中最重要的酶,TCA循环阶段,琥珀酸脱氢酶的铁硫聚簇,FAD和琥珀酸脱氢酶的共价结合,丙二酸 琥珀酸（丁二酸,Succinate is oxidized to fumarate (延胡索酸,1. The enzyme catalyzes stereo

14、specific dehydrogenation 2. Malonate (丙二酸) is the inhibitor of the reaction,该酶具有严格的立体专一性，即只生成反式延胡索酸； 与琥珀酸结构类似的化合物如丙二酸、戌二酸等是该酶的竞争性抑制剂,延胡索酸,苹果酸,Fumarate is hydrated to L-malate (苹果酸,延胡索酸酶,Hydration(水合,该酶具有严格的立体专一性，即只生成L-苹果酸,or,Oxaloacetate (草酰乙酸) is regenerated by the oxidation of L-malate (苹果酸,The st

15、andard free energy of this reaction is very positive. The oxidation of malate is driven by the efficient removal of the products,苹果酸脱氢酶,TCA循环阶段,苹果酸脱氢酶的结构,The last three reactions of the of the TCA cycle are similar to fatty acid degradation and synthesis, and some amino acid degradations. oxidation

16、hydration oxidation,A methylene group (CH2) is converted to a carbonyl group (C=O,The last three reactions of the of the TCA cycle,1. 总反应式: CH3-C-SCoA + 3NAD+ + FAD + GDP + Pi 2CO2 + 3NADHH+ + 1FADH2 + 1GTP + CoA-SH,2. There are two carbon atoms in the form of acetyl-SCoA enters every cycle; and the

17、re are two decarboxylations take place in each cycle; 乙酰辅酶A中的二碳乙酰基进入TCA循环；在循环中发生两次脱羧反应. Notice: the 2 carbon atoms entering the cycle are not the same as those leaving the cycle,Survey of TCA cycle,3. There are four dehydrogenation reactions take place in each cycle; 在循环中发生四次脱氢反应(3NADH,1FADH2,4. The

18、re is a substrate level phosphoralation; 循环中发生一次底物水平磷酸化,Survey of TCA cycle,5. The reactions catalyzed by aconitase and fumarase is asymmetry reactions; 顺乌头酸酶和延胡索酸酶等催化非对称反应 6. All of the intermediary metabolisms are not synthesis and decomposition net. 所有中间代谢产物在代谢中没有净合成或净降解,Survey of TCA cycle,Acety

19、l in acetyl-S-CoA was oxydized: 化学氧化释放的自由能：874.9 kJ/mol 生物氧化释放的自由能：520.9 kJ/mol 用于ATP合成的自由能：354 kJ/mol,The energetic of the aerobic decomposition of glucose,2. Hydrations in TCA-cycle,The third H2O entered in TCA cycle,Citryl-CoA+H2O succinyl-CoA + H2O Fumarate+H2O,Citrate+ CoA-SH Succidate + CoA-SH

20、 Matalte,H2O,H2O,The energetic of the aerobic decomposition of glucose,H2O,3. Dehydrogenations in oxydation of glucose,The energetic of the aerobic decomposition of glucose,4. Caculation of energy,In EMP,ATP 4 - 2,Numbers of ATP,2NADHH+ 2 2.5,In formation of acetyl-CoA,2NADHH+ 2 2.5,In TCA cycle,3NA

21、DHH+ 2 3 2.5 2,FADH2 2 1 1.5 2,GTP 2 1 1 2,Summery: 32,FADH2,2 1.5,30,The energetic of the aerobic decomposition of glucose,The reaction of TCA cycle,1. All organic chemicals can be completely oxidized to CO2 and H2O via a cycled pathway; 所有的有机化合物都可以通过TCA环彻底降解为CO2 和 H2O ; 2. Plenty energy can be pro

22、vided accompany the cycled pathway; 伴随着这一循环途径，大量的自由能被转换,Significance of TCA,3. The intermediates in the cycle are important sources for biosynthetic precursors; TCA环中的中间代谢产物是重要的生物合成的前体 4. The cycle is an amphibolic pathway, and is a metabolic hinge of all biochemicals TCA环是一个分解合成无定向途径，是所有生物化合物的代谢枢纽。

23、分解产能,合成前体,Significance of TCA,In addition to generating reducing potential, the TCA cycle also provides molecules for biosynthetic pathways. Also, the TCA cycle is an entry point for degradative pathways such as amino acid catabolism. Thus, the TCA cycle is highly regulated,TCA Cycle Regulation (TCA

24、循环的调节,Regulation of the PDH Complex,High concentrations of the reaction products of the PDH complex inhibit activity: NADH Acetyl CoA This makes sense because it indicates the cell is rich in energy,Regulation of the PDH Complex,The PDH complex is covalently modified by phosphorylation. Phosphorylation inactivates the complex. Dephosphorylation activates the complex,When the energy charge is high the PDH kinase is