【正文】 UMP C O HN C CCH3 CH N O dR539。P dTMP synthase FH2 N5,N10 methylene FH4 FH4 DHFR NADPH+H+ NADP+ dTMP kinase dTDP kinase ADP dTTP ATP ADP dTMP ATP Thymidylate synthase methylates dUMP at 5position to make dTMP N5,N10methylene THF is 1C donor Synthesis of the Thymine Nucleotides The ribonucleotide reductase, An (R1)2(R2)2 type enzyme , has R1 (86 kD) and R2 ( kD) two subunits E. coli Ribonucleotide Reductase Regulates the level of cellular dNTPs 葉酸 和四氫葉酸 （ FH4） 葉酸 四氫葉酸 H H 10 5 N5， N10CH2FH4 N5CHOFH4 CH2 CHO 核苷酸合成的其他反應(yīng) AMP激酶 核苷二磷酸 激酶 脫氧核糖基轉(zhuǎn)移酶 有 4種針對(duì)不同 NMP的激酶 此酶特異性不強(qiáng) 此酶可以使堿基與脫氧核苷上的堿基交換 ? 嘌呤類似物（ 6巰基嘌呤）：可抑制 AMP、 GMP的生成 ? 谷胺酰胺類似物（氮雜絲氨酸）：可抑制 IMP的合成中有谷胺酰胺參與的反應(yīng) ? 葉酸類似物（氨基蝶呤、氨甲喋呤）：可抑制IMP合成中有四氫葉酸參與的反應(yīng) ?臨床上幾種治癌藥物的作用機(jī)理 嘌呤核苷酸的抗代謝物 嘌呤類似物 : 8氮雜鳥嘌呤 N OH N N N H N SH N N N H N SH N N N H H2N N OH N N N H N 6巰基鳥嘌呤 次黃嘌呤 6巰基嘌呤 （ 6MP） 氨基酸類似物 抑制有谷氨酰胺參與的反應(yīng) H2N—C—CH2—CH2—CH—COOH O NH2 N+ —N—CH2—C—O—CH2—CH—COOH O NH2 N+ —N—CH2—C—CH2—CH2—CH—COOH O NH2 谷氨酰胺 氮雜絲氨酸（重氮乙酰絲氨酸） 6重氮 5氧正亮氨酸 葉酸類似物 抑制有一碳單位參與的反應(yīng) R=H，氨喋呤 R=CH3， 氨甲喋呤 N NH2 N N N H H2N —CH2—N— R —C—N—CH O CH2 CH2 COOH COOH H N N C H —CH2—N— H —C—N—CH O CH2 CH2 COOH COOH H H2N N OH N H 5,6,7,8四氫葉酸 核苷酸的抗代謝物 嘌呤類似物 氨基酸類似物 葉酸類似物 6巰基嘌呤 6巰基鳥嘌呤 8氮雜鳥嘌呤等 氮雜絲氨酸 等 氨蝶呤 氨甲蝶呤 等 次黃嘌呤 (H) 6巰基嘌呤 (6MP) 甲酰甘氨酰 胺核苷酸 （ FGAR） PRPP 谷氨酰胺 （ Gln） = PRA 甘氨酰胺 核苷酸 （ GAR） = = 甲酰甘氨 脒核苷酸 （ FGAM） 5氨基異咪唑 4甲酰胺核苷酸 （ AICAR） = 5甲酰胺基咪唑 4甲酰胺核苷酸 （ FAICAR） IMP 次黃嘌呤 （ H） PRPP PPi = AMP PRPP PPi = 腺嘌呤（ A） GMP = PRPP PPi 鳥嘌呤 (G) 6MP 6MP 6MP 6MP 6MP 6MP 氮雜絲氨酸 氮雜絲氨酸 氮雜絲氨酸 MTX（氨甲喋呤） MTX （氨甲喋呤） Analogs of pymidines /pymidine nucleosides: 5氟尿嘧啶 5Fu 阿糖胞苷 Cytarabine 環(huán)胞苷 Cyclocytidine N N H O O F H HOH2C H H HO OH H H O C C C N N C O NH2 HOH2C H H OH H H O C C C N N C NHHCl O Inhibitors of pymidines synthesis are cancer drugs UMP UTP CTP CDP dCDP UDP dUDP dUMP dTMP 氮雜絲氨酸 阿糖胞苷 氨甲碟呤 氮雜絲氨酸 Deoxyribonucleotide Formation ? Purine/Pyrimidine degradation are the same for ribonucleotides and deoxyribonucleotides ? Biosynthetic pathways are only for ribonucleotide production ? Deoxyribonucleotides are synthesized from corresponding ribonucleotides Formation of Deoxyribonucleotides ? Reduction of 2’ carbon done via a free radical mechanism catalyzed by ―Ribonucleotide Reductases‖ – E. coli RNR reduces ribonucleoside diphosphates (NDPs) to deoxyribonucleoside diphosphates (dNDPs) ? Two subunits: R1 and R2 – A Heterotetramer: (R1)2 and (R2)2 in vitro RIBONUCLEOTIDE REDUCTASE ? R1 SUBUNIT – Three allosteric sites ? Specificity Site ? Hexamerization site ? Activity Site – Five redoxactive –SH groups from cysteines ? R2 SUBUNIT – Tyr 122 radical – Binuclear Fe(III) plex Mechanism of Ribonucleotide Reductase Reaction ? Free Radical ? Involvement of multiple –SH groups ? RR is left with a disulfide group that must be reduced to return to the original enzyme Thioredoxin （硫氧還蛋白） ? Physiologic reducing agent of RNR ? Cys pair can swap H atoms with disulfide formed regenerate original enzyme – Thioredoxin gets oxidized to disulfide Oxidized Thioredoxin gets reduced by thioredoxin reductase mediated by NADPH (final electron acceptor) Thymine Formation ? Formed by methylating deoxyuridine monophosphate (dUMP) ? UTP is needed for RNA production, but dUTP not needed for DNA – If dUTP produced excessively, would cause substitution errors (dUTP for dTTP) ? dUTP hydrolyzed by dUTPase (dUTP diphosphohydrolase) to dUMP ?methylated at C5 to form dTMP ?rephosphorylate to form dTTP Tetrahydrofolate (THF) ? Methylation of dUMP catalyzed by thymidylate synthase – Cofactor: N5,N10methylene THF（ 甲叉四氫葉酸 ）oxidized to dihydrofolate ? THF Regeneration: DHF + NADPH + H+ ? THF + NADP+ (enzyme: dihydrofolate reductase) THF + Serine ? N5,N10methyleneTHF + Glycine (enzyme: serine hydroxymethyl transferase) dUMP dTMP NADPH + H+ NADP+ SERINE GLYCINE REGENERATION OF N5,N10 METHYLENETETRAHYDROFOLATE DHF N5,N10 – METHYLENETHF THF dihydrofolate reductase serine hydroxymethyl transferase thymidylate synthase AntiFolate Drugs ? Cancer cells consume dTMP quickly for DNA replication – Interfere with thymidylate synthase rxn to decrease dTMP production ? (fluorodeoxyuridylate – irreversible inhibitor) – also affects rapidly growing normal cells (hair follicles, bone marrow, immune system, intestinal mucosa) ? Dihydrofolate reductase step can be stopped petitively (DHF analogs) – AntiFolates: Aminopterin（氨蝶呤） , methotrexate（甲氨蝶呤） , trimethoprim （甲氧芐啶） 偶氮絲氨酸 阿西維辛 核苷酸分解代謝 ? 嘌呤堿的分解首先是在各種脫氨酶的作用下水解脫去氨基。 ? 脫氨反應(yīng)也可以在核苷或核苷酸的水平上進(jìn)行，在動(dòng)物組織中腺嘌呤脫氨酶的含量極少，而腺嘌呤核苷脫氨酶和腺嘌呤核苷酸脫氨酶的活性極高。 ? All purine degradation leads to uric acid (but it might not stop there) Gout ? Impaired excretion or overproduction of uric acid ? Uric acid crystals precipitate into joints (Gouty Arthritis), kidneys, ureters (stones) ? Xanthine oxidase （黃嘌呤氧化酶） inhibitors inhibit production of uric acid, and treat gout ? Allopurinol treatment – hypoxanthine analog that binds to Xanthine Oxidase to decrease uric acid production ALLOPURINOL IS A XANTHINE OXIDASE INHIBITOR A SUBSTRATE ANALOG IS CONVERTED TO AN INHIBITOR, IN THIS CASE A “SUICIDEINHIBITOR” 高嘌呤食物：豆苗、黃豆芽、蘆筍、香菇、紫菜、動(dòng)物內(nèi)臟、魚類 ? 腺苷脫氨酶缺乏癥 Aadenosine deaminase deficiency (ADA) ：一種嚴(yán)重