生物化学教学课件：复习课

1、Protein breakdown starts at stomachOptimal pH1.52.5，Low specificityProduce small peptide and free amino acids pepsinogen胃蛋白酶原+ Small PeptideHCl、PepsinPepsinBreakdown of dietary proteins Enterokinase肠激酶 : produced by cells of the duodenum du:di:nm （十二指肠）Synthesis of the enzymes as inactive precursors

2、-zymogen(酶原或前酶）:1 Protects the exocrine cells from destructive proteolytic attack. 2 Transportation3 Allow RegulationElaborate mechanism for getting active digestive enzymes.Protein Digestion Proteolytic enzymes of pancreatic juiceEndopeptidasesExopeptidasesTrypsin : Arg, Lys (C)Chymotrypsin : Tyr,

3、Trp, Phe, Met, Leu (C)Elastase: Ala, Gly, Ser (C)carboxypeptidaseaminopeptidaseCleavage sites and substrates of proteasesMARTKQTARKSTGGKAPRKQLATKAARKSAPSTGTwo major systems for the turnover of cellular proteins:Lysosomes and Ubiquitin-mediated protein degradationProtein Degradation in Cell1 Lysosoma

4、l hydrolasesexhibit acidic pH optimal.2 Lost activity if leaked into cytosol.3 Cathepsin 组织蛋白酶家族Low specificityLysosomal Degradation PathwayProteinsA. EndocytosedB. Cytosolic C. Organelles by autophagy溶酶体贮积病：基因突变致溶酶体酸性水解酶缺陷痛风：溶酶体破裂Ubiquitination Ub activating enzyme(Uba) Ub conjugating enzyme(Ubc) E

5、3- Ub LigaseWhat is Ubiquitin and Ubiquitylation?Human Ub: MQIFKVTLTGKTITLEVEPNDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLADYNIQKESTLHLVLRLRGGYeast Ub: MQIFKVTLTGKTITLEVESSDTIDNVKSKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNIQKESTLHLVLRLRGG肽键和异肽键substrateUbMQIFKVTLTGKTITLEVEPNDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLADYNIQ

6、KESTLHLVLRLRGGUbiquitin pathwayE1-S-UbE2-S-UbE3substratesUbMono-ubiquitinationChange：protein conformation,Localization, interactionPolyubiquitinationUbUbUbUbsubstratesProteosomeDegradationE1- Ub activating enzyme(Uba)Uses ATPAMP+PPi to form thioester bond between its sulfhydryl group and the termina

7、l carboxylate group of UBE2- Ub conjugating enzyme(Ubc):Accepts activated UBE3- Ub Ligase: Catalyzes transfer of UB from E2 to -amino group of lysine on target protein forming an isopeptide bond异肽键ATP蛋白质泛素化降解途径7要素Amino acid metabolism Deamination（脱氨） Transamination（转氨作用） Urea cycle（尿素循环） Pyruvate gr

8、oup a Ketoglutarate group Succinyl-CoA group Oxaloacetate group Fumarate / oxaloacetate group Alanine / acetoacetate group Acetyl-CoA / acetoacetate group DecarboxylationTransamination(转氨作用）ALT: Alanine aminotransferase (in liver) 丙氨酸氨基转移酶，also called glutamate-pyruvate transaminase（谷丙转氨酶）, GPTAST:

9、Aspartate aminotransferase (in heart) ，also called glutamate-oxaloacetate transaminase（谷草转氨酶）, GOTTwo important transaminases:Transamination-needs coenzyme pyridoxal phosphate（磷酸吡哆醛， B6）1 Covalent ligated to enzyme2 Interaction with enzyme3 Schiffs baseVitamine B6亚胺(2) Oxidative deaminationGlutamate

10、 Releases Its Amino Group as Ammonia in the Liver1 In hepatocytes mitochondria, 2 Glutamate undergoes oxidative deamination 3 Catalyzed by L-glutamate dehydrogenase4 Produces free ammonia5 Need NAD and Produces NADH 6 Required for Urea Cycle7 The enzyme is strictly regulated by energy (ADP, ATP)Meta

11、bolism of Ammoniamost vertebratesbirds & reptilesfish & other aquatic vertebrates1 Glucose-Alanine CycleAlanine Transports Ammonia fromSkeletal Muscles to the Liver1 Glucose Alanine cycle2 Transportation of ammonia by Gln 丙氨酸：NeutralNon-polarityNon-toxicEasily transported through membrane2. Transpor

12、tation of ammonia by GlnExcess ammonia in tissues is added to glutamate to form glutamine, catalyzed by glutamine synthetase(谷氨酰胺合成酶） After transport in the bloodstream,the glutamine enters the liver and NH4 is liberated in mitochondria by the enzyme glutaminase（谷氨氨酰胺酶）Formation of urea1. Site: live

13、r (mitochondria and cytosol)2. Process - ornithine cycleUrea Is Produced from Ammonia in Five Enzymatic Steps(氨、瓜、精琥、精、尿）Hans Creb1 The 2 N of urea : NH3 and Asp.2 Starts/ends up with: ornithine3 NH3, CO2 form caramoyl phosphate,4 Citrulline formation,5 Incorporation of Asp.6 Energy consuming: One u

14、rea 3ATP, 4 P7 Needs 5 enzymes：carbamoyl phosphate synthetase ornithine carbamoyl transferase, OCTArgininosuccinate SynthetaseArgininosuccinaseArginase8 Rate limiting :1st step9 mitochondrianl vs cytosolUrea formation:Regulation of urea synthesisActivation of the urea cycle by N-acetyl-glutamateN-ac

15、etyl-glutamate synthetase IRegulation of urea synthesisEnergy consuming: one urea will costs 4 PCarbamoylphasphate synthetase ISubstrate activation/inductionResponsive to concentration of ammonia /amino acid in liverProtein-rich dietary Starvation Five final products of the backbone of amino acids:

16、connected to TCAAcetyl-CoAa-ketoglutarateSuccinyl-CoAFumarateOxaloacetateTetrahydrofolic acid (FH4) and one carbon unitOne carbon units are produced in catablite of AA and carried by FH4. One carbon units are substance for synthesis of nucleic acid.In great need of new-born蝶啶对氨基苯甲酸GlutamatePurine ri

17、ng甲基受体接受甲基甲硫氨酸腺苷甲硫氨酸AdoMet, SAMMethylatedacceptor高半胱氨酸FH4 provides methyl group for methionine, which servers as major methyl-group donor for many reactions, Branched Chain amino acid(支链氨基酸, BCAA)的起始途径是相同的： aminotransferases 同工酶Branched keto acidsDehydrogenase支链氨基酸分解代谢酶缺乏并不多见，一旦发生可造成酸中毒；支链酮酸脱氢酶复合体活性

18、缺失导致枫树蜜汁尿病：尿液具有特殊的枫树蜜汁气味，伴随酸中毒，智力迟缓和短寿。限制膳食治疗！PhenylKetonuria苯丙酮尿症Phenylalanine HydroxylaseTyrosine AminotransferaseParahydroxyphenyle pyruvate Di-oxygenase (ascorbate Vc)Homogentisate DioxygenaseMaleyl Acetoacetate IsomaeraseFumaryl AcetoacetaseAlanineAsparagineAspartateCysteineGlutamateGlutamineGl

19、ycineProlineSerineTyrosineArginine*HistidineIsoleucineLeucineLysineMethioninePhenylalanineThreonineTryptophanValineEssential and Nonessential Amino Acids for Humans Essential Nonessential*Arg is essential in infants and growing children but not in adults. Most bacteria and plants can synthesize all

20、20 amino acids.AAs that an organism cannot synthesize under a given set of conditions are called “essential.” Those that can be synthesized in adequate amounts are “nonessential.”23 Humans can synthesize 10 of the 20 common amino acidsA, B, C 三种酶催化三步反应；每种酶有2-3个同工酶；其中有一个酶是非变构酶；当Met高的时候，A2 and B1的合成被抑

21、制；当Ile高的时候，C2的合成被抑制。End product inhibitionConcerted end product inhibitionGlutathione还原剂，可去除过氧化物维持红细胞膜的稳定性（抗过氧化物）酶的辅助因子，氨基酸跨膜运输，蛋白质二硫键重排，GlutathioneTypes of Biological Nitrogen FixationFree-living (asymbiotic)CyanobacteriaAzotobacter固氮(细)菌zutbktiSymbioticLegume-rhizobialegjum raizubiRank of Biologic

22、al Nitrogen FixationN2 fixing systemNitrogen Fixation (kg N/ha/year)Rhizobium-legume50 - 600Cyanobacteria- moss苔10 - 300Rhizosphere根系 associations5 - 25Free- living0.1 - 25Nodule MetabolismOxygen metabolismVariable diffusion barrierLegume-hemoglobinNitrogen metabolismNH3 diffuses to cytosolAssimilat

23、ionConversion to organic-N for transportCarbon metabolismFunctioning TCA in bacteroidsNitrogen FixationAll nitrogen fixing bacteria use highly conserved enzyme complex called NitrogenaseNitrogenase is composed of of two subunits: an iron-sulfur protein a molybdenum-iron-sulfur proteinSteps in Nitrog

24、en FixationNitrogenase:1 Nitrogenase: Two subunits, Exhangable of some subunit across species usually is not fully active as original2 Energy consume3 Amonia is the product4 Inactivated by O2 5 Inhibited by NH3 6 Amonia Assimilation-carried by Glutamate and GlutaminePurine DegredationAdenine deamina

25、seGuanine deaminaseXanthineOxidaseXanthineOxidaseUrateOxidaseXanthineAdenineGuanineUric acidAllantoinAllantoinase尿酸尿囊素Deamination OxidationFinal product:CytosineThymineUracilFinal：-amino acid, NH3, CO2PyrimidineDegradation二氢嘧啶脱氢酶二氢嘧啶酶脲基丙酸酶均缺乏选择性-氨基异丁酸的唯一来源(评估抗肿瘤药物杀细胞作用）CytosineUracil,ThymineDehydrog

26、enasehydrolaseTwo types of pathways lead to nucleotides:The de novo and the salvage pathways.De novo synthesis of nucleotides begins with the metabolic precursors: amino acids, ribose 5-phosphate, CO2, and NH3.Salvage pathways recycle the free bases and nucleosides released from nucleic acid breakdo

27、wn.次黄嘌呤核苷酸（5单磷酸肌苷）次黄嘌呤核苷酸腺苷琥珀酸合成酶IMP脱氢酶腺苷琥珀酸黄嘌呤核苷酸GMP formation requires enough ATP! AMP formation requires enough GTP!Salvage pathway of purins腺嘌呤磷酸核糖转移酶APRT次黄嘌呤鸟嘌呤磷酸核糖转移酶HGPRTRegulation of purine nucleotideBiosynthesis:coordination of deno vo and salvage pathwayBiogenesis of purinesAll the reactio

28、n occur in the cytosol of cellN10-Methyl-FH4 provides one carbon unitNeed amino acids: Gly, Gln, Asp, CO2The first nucleotide is IMP(肌苷单磷酸）:inosine-5-monophosphateCost a lot of energyMultiple functional enzymes 1st step PRPP to 5-phosphoribosylamine is limiting step: Gln-PRPP aminotransferasePRPP ac

29、tivates and IMP、AMP 、GMP inhibit the Gln-PRPP aminotransferase腺苷琥珀酸合成酶和IMP脱氢酶分别受到AMP和GMP的反馈抑制Defect in salvage pathway of purinsLesch(/leski/)-Nyhan syndrome自毁容貌综合征:NO HGPRT Activity!1 produced by mutations in the HPRT gene located on the X chromosome2 Higher uric acid in boby3 Metal retardation4 Se

30、lf-mutilationThe GoutOverproduction of uric acidUric acid crystals precipitate into joints (Gouty Arthritis痛风性关节炎), kidneys, ureters (stones结石)Adenine deaminaseGuanine deaminaseXanthineOxidaseAdenineSalvage pathway deficiency or Cancer treatment, Allopurinol :hypoxanthine analog Xanthine oxidase inhibitors binds to Xanthine Oxidase inhibit production of uric acid别嘌呤醇次黄嘌呤Pyrimidine biosynthesisCarbamoylphosphateAspartate1 由氨甲酰磷酸和天冬氨酸2 氨甲酰磷酸由氨甲酰磷酸合成酶II催化3 氨甲酰磷酸合成酶II存在于细胞质中不同于氨甲酰磷酸合成酶I存在于线粒体中4 先合成嘧啶环UMP合成酶缺陷：遗传性乳清