生物化学_Glycolysis

18.Glycolysis

basic
- Carried out in the cytosol of cells, it is basically an anaerobic 厌氧 process
- its principal steps occur with no oxygen
- provide precursor molecules for aerobic catabolic pathways 有氧代谢过程
- Compartmentalization of glycolysis, the citric acid cycle, and oxidative phosphorylation.
- 图
two phase
- first phase
  - basic
    - glucose → two molecules of glyceraldehyde-3-phosphate (G-3-P) 3-磷酸甘油醛
    - consume 2 ATP
  - Reaction 1 - Phosphorylation of Glucose → G-6-P (-ATP) 调控步骤
    The First Priming Reaction
    - glucose → Glucose-6-Phosphate
    - Cellular Advantages 对细胞有什么好处
      - phosphorylation keeps the substrate in the cell, G-6-P带负电出不去
      - keeps the intracellular concentration of glucose low, 可以让更多glucose扩散进来
      - because regulatory control can be imposed only on reactions not at equilibrium (∆G=0), the favorable thermodynamics of this first reaction makes it an important site for regulation 这步反应可以用于调节
    - 酶 - Hexokinase 己糖激酶 or Glucokinase 葡(萄)糖激酶
      - Hexokinase 己糖激酶
        
        In most animal, plant, and microbial cells, the enzyme that phosphorylates glucose is hexokinase
        
        can phosphorylate a variety of hexose sugars, including glucose, mannose 甘露糖, and fructose 果糖
        
        Mg2+ is required for this reaction, the true substrate for the hexokinase reaction is MgATP2-
        
        The Km for glucose is ~0.1 mM, and the enzyme thus work efficiently at normal blood glucose levels of 4 mM
        
        regulation
        
        Different body tissues possess different isozymes同工酶 of hexokinase , each exhibiting somewhat different kinetic properties
        
        负反馈调节
        
        The animal enzyme is allosterically inhibited by the product, G-6-P
        
        被产物变构抑制
        
        levels of G-6-P inhibit hexokinase activity until consumption by glycolysis lowers its concentration
        
        The hexokinase reaction is one of three points in the glycolysis pathway that are regulated
      - Glucokinase 葡(萄)糖激酶
        
        Liver contains an enzyme called glucokinase
        
        highly specific for D-glucose
        自然界中存在的葡萄糖
        
        has a much higher Km for glucose, ~10.0 mM, works only when liver glucose levels are high, and store G-6-P in liver as glycogen
        
        not product-inhibited
        
        is an inducible enzyme诱导酶, the amount present in the liver is controlled by insulin 胰岛素
        环境中有信号诱导酶才会大量合成, Patients with type 2 diabetes 糖尿病 produce insufficient insulin
    - Glucose-6-phosphate is the branch point for several metabolic pathways
      - 图
  - Reaction 2 - the Isomerization 异构化 of G-6-P → F-6-P
    - Glucose-6-Phosphate → Fructose-6-Phosphate
    - 具体机制
      - 醛酮互变异构
      - 图
      - opening of the pyranose 吡喃糖ring (Step A)
      - proton abstraction leading to enediol烯二醇formation (Step B)
      - and proton addition to the double bond, followed by ring closure (Step C)
      - The furanose 呋喃糖 is formed by attack of the C-5 hydroxyl on the carbonyl group
    - 作用
      - First, the next step in glycolysis is phosphorylation at C-1, and the hemiacetal半缩醛-OH of glucose would be more difficult to phosphorylate than a simple primary hydroxyl 羟基.
      - 第一，糖酵解的下一步是在 C-1 位发生磷酸化，而葡萄糖的半缩醛羟基比简单的伯羟基更难发生磷酸化
      - Second, the isomerization to fructose (with a carbonyl group at position 2 in the linear form) activates carbon C-3 for cleavage in the fourth step of glycolysis
      - 第二，异构化为果糖（其线性形式中 C-2 位含羰基）可激活 C-3 位碳，以便在糖酵解第四步中发生断裂
    - 酶 - phosphoglucose isomerase 磷酸葡萄糖异构酶
      - phosphoglucose isomerase 磷酸葡糖异构酶, also known as glucose phosphate isomerase
      - requires Mg2+, highly specific for glucose-6-phosphate
      - ∆G =1.67kJ/moL namely, 0.4 kcal/mol, means that the reaction operates near equilibrium in the cell and is readily reversible 在细胞内接近平衡, 容易逆向发生
  - Reaction 3 - F-6-P → FBP (-ATP) 调控步骤
    The Second Priming Reaction
    - Fructose-6-Phosphate → Frutose-1,6-BiPhosphate
    - At pH 7 and 37°C
      - the phosphofructokinase reaction equilibrium lies far to the right
      - commits 指派the cell to metabolizing glucose rather than converting it to another sugar or storing it
    - 酶 - phosphofructokinase 磷酸果糖激酶
      - Phosphofructokinase with ADP shown in white and fructose-6-P in red
      - most important site of regulation in the glycolytic pathway
      - regulation - allosteric
        
        ↓ ATP
        
        ATP is an allosteric inhibitor of this enzyme
        
        adenylate kinase腺苷酸激酶会催化以下反应
        
        ATP and AMP levels represent the cellular energy status
        
        ↓ citrate 柠檬酸盐
        
        Glycolysis and TCA are coupled via phosphofructokinase
        
        citrate 柠檬酸盐, an intermediate in the citric acid cycle, is an allosteric inhibitor of phosphofructokinase
        
        When the citric acid cycle reaches saturation, glycolysis slows down
        
        ↑ AMP
        
        AMP reverses the inhibition due to ATP
        
        ↑ β-D-fructose-2,6-bisphosphate
        
        三个功能
        
        increases the affinity of phosphofructokinase for the substrate fructose-6-phosphate
        
        decreasing the inhibitory effects of ATP
        
        inhibiting fructose-1,6bisphosphatase, the enzyme that catalyzes this reaction in the opposite direction
        
        increases the net flow of glucose through glycolysis by stimulating phosphofructokinase
        
        这个增强剂与这步反应的产物对比
        
        被PFK-2 & F-2,6-BPase调控
  - Reaction 4 - Cleavage of FBP → DHAP + G-3-P
    - Fructose-1,6-bisP → DiHydroxyAcetone Phosphate ＋ Glyceraldehyde-3-Phosphate
    - cleaves fructose-1,6-bisphosphate between the C-3 and C-4 carbons to yield two triose phosphates 磷酸丙糖
      - dihydroxyacetone phosphate (DHAP) 磷酸二羟基丙酮
      - glyceraldehyde-3-phosphate (G-3-P) 3-磷酸甘油醛
    - 酶 - Fructose bisphosphate aldolase 果糖二磷酸醛缩酶
      - equilibrium constant of approximately 10^-4 M, and a corresponding ∆G° of 23.9 kJ/mol.
      - 表面数据这个反应很难发生, 但是是一个分子产生两个分子, 受到浓度影响大
      - The value of ∆ G in erythrocytes 红细胞 is actually -0.23 kJ/mol.
      - At physiological concentrations, the reaction is essentially at equilibrium
  - Reaction 5 - DHAP ⇄ G-3-P
    - DiHydroxyAcetone Phosphate ⇄ Glyceraldehyde-3-Phosphate
    - 这步和上一步加起来相当于FBP → 2G-3-P
    - 只有G-3-P能进入glycolysis二阶段, 所以要把DHAP变成G-3-P
    - 酶 - Triose Phosphate Isomerase 磷酸丙糖异构酶
  - energy
    - Although the last two steps of the pathway are energetically unfavorable, the overall five-step reaction sequence has a net ∆G°of +2.2 kJ/mol (Keq≈0.43).
    - It is the free energy of hydrolysis from the two priming molecules of ATP that brings the overall equilibrium constant close to 1 under standard-state conditions.
    - The net ∆ G under cellular conditions is quite negative (-53.4 kJ/mol in erythrocytes)
- second phase
  - basic
    - two molecules of glyceraldehyde-3-phosphate → two molecules of pyruvate 丙酮酸
    - produce 4 ATP
  - Reaction 6 - G-3-P → 1,3-BPG (-2P +2NADH)
    - G-3-P → 1,3-BPG
    - makes a high-energy phosphate in preparation for ATP synthesis
    - 酶 - Glyceraldehyde-3-Phosphate Dehydrogenase 脱氢酶
      - Although the oxidation of an aldehyde to a carboxylic acid is a highly exergonic放能 reaction
      - the overall reaction involves both formation of a carboxylic-phosphoric anhydride and the reduction of NAD to NADH
      - therefore slightly endergonic吸能的 at standard state, with a ∆G of +6.30 kJ/mol
  - Reaction 7 - 1,3-BPG → 3-PG (+2ATP)
    - 1,3-BPG → 3-PG
    - 酶 - Phosphoglycerate Kinase 磷酸甘油酸激酶
      - Mg2+ ion is required for activity
      - exergonic 放能
      - In fact, the aldolase 醛缩酶 and triose phosphate isomerase 磷酸三糖异构酶 are also pulled forward by phosphoglycerate kinase
        这步放能拉着前面两步能量不利的往前反应
  - Reaction 8 - phosphoryl transfer
    - 3-PhosphoGlycerate → 2-PhosphoGlycerate
    - 酶 - Phosphoglycerate mutase 磷酸甘油酸变位酶
      - The term mutase 变位酶 is applied to enzymes that catalyze migration of a functional group within a substrate molecule
      - The free energy change for this reaction is very small under cellular conditions
  - Reaction 9 - Dehydration 2-PG → PEP (-H2O)
    - 2-PhosphoGlycerate → PhosphoEnolPyruvate 磷酸烯醇丙酮酸
    - makes a high-energy phosphate in preparation for ATP synthesis
    - 酶 - Enolase 烯醇酶
      - The ∆G° for this reaction is relatively small at 1.8 kJ/mol (Keq 0.5)
      - under cellular conditions, ∆G is very close to zero
  - Reaction 10 - PEP → pyruvate (+2ATP) 调控步骤
    - PhosphoEnolPyruvate → pyruvate
    - 酶 - Pyruvate Kinase 丙酮酸激酶
      - The reaction requires Mg2+ ion and is stimulated by K+ and certain other monovalent cations单价阳离子
      - Keq at 25°C is 3.63×10^5, equilibrium far right
      - The large negative G of this reaction makes pyruvate kinase a target site for regulation of glycolysis 适合调控
      - regulation - allosteric
        
        ↑ AMP
        
        ↑ fructose-1,6-bisphosphate
        
        ↓ ATP
        
        ↓ acetyl-CoA
        
        ↓ alanine丙氨酸
        
        Furthermore, liver pyruvate kinase is regulated by covalent modification
The Products of Glycolysis
- ATP
- NADH
  - NADH must be recycled to NAD+, lest以免 NAD+ become limiting in glycolysis.
  - NADH can be recycled by both aerobic需氧的 and anaerobic厌氧的 paths, either of which results in further metabolism of pyruvate.
- pyruvate 丙酮酸
after glycolysis
- Aerobic 有氧
  - pyruvate sent into the citric acid cycle (TCA)
  - it is oxidized to CO2 with the production of additional NADH (and FADH2).
  - the NADH produced in glycolysis and the citric acid cycle is reoxidized to NAD+ in the mitochondrial electron transport chain
  - 图
- Anaerobic 无氧
  - fermentation 发酵
    - pyruvate → lactate/ethanol都算fermentation
    - the production of ATP energy by reaction pathways in which organic molecules function as donors and acceptors of electrons
    - In either case, reduction of pyruvate provides a way of reoxidizing the NADH produced in the glyceraldehyde-3-phosphate dehydrogenase reaction of glycolysis
  - pyruvate → lactate 乳酸
    in microorganisms(other than yeast) and animals
    - NAD is regenerated in the lactate dehydrogenase reaction
    - produce the taste of sour milk characteristic taste and fragrance of sauerkraut 泡菜, which in reality is fermented cabbage
    - Lactate Accumulates Under Anaerobic Conditions in Animal Tissues
      - Pyruvate reduction occurs in tissues that normally experience minimal access to blood flow (e.g. the cornea 角膜) and also in rapidly contracting skeletal muscle
      - In anaerobic muscle tissue, lactate represents the end of glycolysis
      - cramps 抽筋 and muscle soreness 酸痛, fatigue 疲劳 associated with the buildup 堆积 of lactic acid in the muscle
      - Most of this lactate must be carried out of the muscle by the blood and transported to the liver, where it can be resynthesized into glucose in gluconeogenesis
    - 酶 - lactate dehydrogenase 乳酸脱氢酶 LDH
      - Mammalian LDH exists as five different isozymes同工酶
        Enzymes with Slightly Different Subunits, 催化同一种反应但是催化效率与倾向不同
      - depending on the tetrameric 四聚体 association of two different subunits(A&B)
      - A4, A3B, A2B2, AB3, B4
      - their relative affinities for the various substrates and their sensitivity to inhibition by product are different
      - Different tissues express different isozyme forms
        
        muscle
        
        anaerobic
        
        needs LDH to regenerate NAD from NADH so glycolysis can continue
        
        The muscle LDH isozyme (A4) works best in the NAD regenerating direction
        
        heart
        
        aerobic 有氧
        
        uses lactate as a fuel, converting it to pyruvate via LDH
        
        using the pyruvate to fuel the citric acid cycle to obtain energy
        
        The heart LDH isozyme (B4) is inhibited by excess pyruvate so the fuel won’t be wasted
        
        图
    - Lactic acid fermentation
  - pyruvate → ethanol + CO2
    in yeast
    - provides a means for regenerating NAD consumed in the glyceraldehyde-3-P dehydrogenase reaction
    - Alcoholic fermentations are the basis for the brewing酿造 of beers and the fermentation of grape sugar in wine making
    - Alcoholic fermentation
- 总结
  - 图
其他
- energy elegance
  - 反应在标准状态以及细胞内的自由能
  - 细胞内反应2和反应4-9 ∆G is very close to zero, so that these reactions operate essentially at equilibrium
  - The driving force for glycolysis lies in the hexokinase (1), phosphofructokinase (3), and pyruvate kinase (10) reactions
  - These reactions are thus the sites of glycolytic regulation
  - gluconeogenesis 糖质新生
    - the biosynthesis of glucose
    - different enzymes are used to carry out reactions 1, 3, and 10 in reverse, effecting the net synthesis of glucose.
    - The maintenance of reactions 2 and 4 through 9 at or near equilibrium permits these reactions (and their respective enzymes) to operate effectively in either the forward or reverse direction
- Other Substrates in Glycolysis
  - Mannose, galactose, fructose, and other simple metabolites can enter the glycolytic pathway through being converted by appropriate enzymes to one of the intermediates of glycolysis.
- Nucleoside 5’-Triphosphates Are Carriers of Chemical Energy
  - ATP, GTP, CTP, UTP and dATP, dGTP, dCTP, dTTP.
    Four NTPs and their dNTPcounterparts are the substrates for the synthesis of the nucleic acids
  - The evolution of metabolism has led to the dedication of one of these four NTPs to each of the major branches of metabolism
  - ATP has been termed the energy currency of the cell.
  - GTP is the major energy source for protein synthesis.
  - CTP is an essential metabolite in phospholipid synthesis.
  - UTP forms activated intermediates with sugars that go on to serve as substrates in the biosynthesis of complex carbohydrates and polysaccharides