Nucleic Acid Metabolism PDF

Nucleic Acids Metabolism DNA & RNA  Nucleic Acid Metabolism There are two aspects Biosynthesis or anabolism Degradation or catabolism Focus Purine metabolism & Regulation Pyrimidine metabolism & Regulation Abnormalities in Nucleic acid Metabolism This...

Nucleic Acids Metabolism DNA & RNA  Nucleic Acid Metabolism There are two aspects Biosynthesis or anabolism Degradation or catabolism Focus Purine metabolism & Regulation Pyrimidine metabolism & Regulation Abnormalities in Nucleic acid Metabolism This metabolism is essential for cellular functions, including replication, transcription, and translation, which are critical for gene expression and cellular function.  Nucleic Acid Metabolism Modern relevance to medicine includes: Cancer Therapy: Targeting nucleic acid metabolism pathways has led to the development of chemotherapeutic agents that inhibit DNA synthesis or repair. Genetic Disorders: Understanding nucleic acid metabolism aids in the diagnosis and treatment of numerous genetic diseases, enabling gene therapy approaches. Antiviral Treatments: Nucleoside analogs are used to treat viral infections by interfering with viral nucleic acid synthesis. Personalized Medicine: Advances in genomics and RNA sequencing help tailor treatments based on individual genetic profiles.  What do they do ? Dictate amino-acid sequence in proteins Give information to chromosomes, which is then passed from parent to offspring  What are they ? The 4th type of macromolecules The chemical link between generations The source of genetic information in chromosomes The central dogma of molecular biology. The central dogma of molecular biology. Nucleic Acids and Heredity  Processes in the transfer of genetic information:  Replication: identical copies of DNA are made  Transcription: genetic messages are read and carried out of the cell nucleus to the ribosomes, where protein synthesis occurs.  Translation: genetic messages are decoded to make proteins.  NUCLEIC ACIDS (DNA and RNA) Notes DNA – Deoxyribonucleic Acid DNA controls all living processes including production of new cells – cell division DNA carries the genetic code – stores and transmits genetic information from one generation to the next Chromosomes are made of DNA DNA is located in the nucleus of the cell  Nucleotides and Nucleosides  Nucleotide =  Nitrogeneous base  Pentose  Phosphate  Nucleoside =  Nitrogeneous base  Pentose  Nucleobase =  Nitrogeneous base  Formation of nucleic acids Nucleotides are linked through phosphate groups and 3’ position of the sugar (3’ – 5’ phosphodiester linkage)  Primary Structure of Nucleic Acids  The primary structure of a nucleic acid is the nucleotide sequence  The nucleotides in nucleic acids are joined by phosphodiester bonds  The 3’-OH group of the sugar in one nucleotide forms an ester bond to the phosphate group on the 5’-carbon of the sugar of the next nucleotide Generalized Structure of DNA Generalized Structure of DNA  Nucleic acid metabolism The Nucleic acid metabolism section will focus on biosynthesis and catabolism of the nucleotides, and the diseases that condition associated with the result of defects in the enzymes of the pathways of nucleotide biosynthesis and catabolism. 1. DE NOVO BIOSYNTHETIC PATHWAYS (building the bases from simple building blocks) 2. SALVAGE PATHWAYS (the reutilization of bases from dietary or catabolic sources)  Nucleic acid metabolism SYNTHESIS OF PURINE RIBONUCLEOTIDES SYNTHESIS OF PURINE RIBONUCLEOTIDES IMP is synthesized through the assembly of a purine base on ribose-5-phosphate. Kinases convert IMP-derived AMP and GMP to ATP and GTP. Purine nucleotide synthesis is regulated by feedback inhibition and feedforward activation. Salvage reactions convert purines to their nucleotide forms. SYNTHESIS OF PURINE RIBONUCLEOTIDES The biosynthesis of purine (A and G) begins with the synthesis of the ribose-phosphate  SYNTHESIS OF PURINE RIBONUCLEOTIDES The major regulatory step in purine biosynthesis is the conversion of PRPP to 5- Phosphoribosyl-1-amine Amidophosphoribosyl transferase is an important regulatory enzyme in purine biosynthesis. It is strongly inhibited by the end products IMP, AMP, and GMP. This type of inhibition is called FEEDBACK INHIBITION.  SYNTHESIS OF PURINE RIBONUCLEOTIDES Several amino acids are utilized in purine biosynthesis, IMP is the precursor for both AMP and GMP, the base is also called hypoxanthine  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP Methotrexate- A competitive inhibitor of dihydrofolate reductase - role in purine & pyrimidine biosynthesis, Is used to treat cancer Pteridine + PABA/ sulpho.. Dihydropteroate synthase Dihydropteroic acid Dihydrofolic acid Dihydrofolate reductase Tetrahydrofolic acid  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES de novo biosynthesis of IMP  SYNTHESIS OF PURINE RIBONUCLEOTIDES Conversion of IMP to AMP and GMP showing feedback inhibition  SYNTHESIS OF PURINE RIBONUCLEOTIDES Conversion of Hypoxanthine to Adenine/Guanine. The common mechanistic theme for the conversion of A and G is the conversion of a carbonyl oxygen to an amino group  Formation of Di & Tri-phosphates The nucleoside monophosphates (AMP & GMP) are converted to the corresponding di & triphosphates , By the transfer of phosphate group from ATP (phosphorylation), catalysed by specific nucleoside monophosphate (NMP) kinases to form nucleoside diphosphates & less specific nucleoside diphosphate (NDP) kinases to form nucleoside triphosphates.  Formation of Deoxyribonucleotides from Ribonucleotides Deoxyribonucleotides(building blocks of DNA) are derived from corresponding ribonucleotides by reduction of 2’-C of the ribose to form 2’- deoxyribose(i.e the 2’-hydroxyl group on the ribose is replaced by a hydrogen). Catalyzed by Ribonucleotide reductase enzyme. Ribonucleotide reductase catalyzes the rate-determining step in biosynthesis of DNA precursor NADPH donates a pair of H-atom via thioredoxin. Ribonucleoside diphosphates or triphosphates are the Substrates. BCH 212  Regulatory Control of Purine Nucleotide Biosynthesis Purine Biosynthesis are precisely regulated by feedback inhibition. Four key enzymes are majorly feedback regulated: PRPP synthetase,Glutamine phosphoribosyl amidotransferase ,adenylosuccinate synthetase & Inosine Monophosphate (IMP) dehydrogenase. 1. PRPP synthetase (enzyme converting Ribose-5-phosphate to Phosphoribosyl pyrophosphate( PRPP)) is negatively feedback inhibited by GDP & also inhibited at a distinct allosteric site by ADP & also regulated by metabolites from other pathways that PRPP is a starting point. 2. Glutamine phosphoribosyl amidotransferase is activated by PRPP & competitively feedback inhibit (inactivated) by AMP ,GMP & IMP. The active Glutamine phosphoribosyl amidotransferase is a monomer but converted to an inactive dimer by binding of the end products(AMP,GMP & IMP). PRPP causes a shift towards the active monomeric form 3. Adenylosuccinate synthetase (Enzyme that converts IMP to adenylosuccinate( an immediate precursor of AMP)) is also feedback competitively inhibited by AMP without affecting the biosynthesis of GMP. 4. IMP dehydrogenase (Enzyme converting IMP into xanthylate/xanthosine monophosphate (XMP)., is feedback competitively inhibited by GMP, without affecting the biosynthesis of AMP.  SYNTHESIS OF PURINE RIBONUCLEOTIDES The regulation of purine biosynthesis is a classic example of negative feedback IMP pathway regulated at first two steps – Feedback inhibition – Feedforward activation Highlights of Purine de-novo synthesis & Regulations SYNTHESIS OF PURINE RIBONUCLEOTIDES  Purine Salvaging  Free purines released in nucleic acid degradation  Can be converted into corresponding nucleotides through salvage pathways  Salvage biosynthesis of Purine Occurs mainly in extrahepatic tissues/cells such as brain, bone marrow & erythrocyte (In animals) & plastide (In plants & fungi). The salvage pathway is especially important in certain tissues such as erythrocytes, polymorph nuclear leukocytes & brain where denovo synthesis of purine nucleotides is not operative as lesser amount of aminotransferase. Occurs by phosphorylation ,with PRPP as the donor of ribose-5- phosphate,but requires less energy than De-novo pathway for purine synthesis. Involves free purines base(A & G) from nucleic acid breakdown or diet being joined with phosphoribosyl pyrophosphate (PRPP) to produce mononucleotides by two enzymes: 1. Adenine phosphoribosyl transferase (APRT):Catalyzes formation of Adenosine monophosphate(AMP) from Adenine and phosphoribosyl pyrophosphate (PRPP). 2. Hypoxanthine-guanine phosphoribosyl transferase (HGPRT): catalyzes the formation of Inosine Monophosphate(IMP) using Hypoxanthine & PRPP.Also catalyze the formation of Guanosine Monophosphate(GMP) from Guanine & PRPP Deficiency of HGPRT leads to Lesch-Nyhan syndrome. Results in failure to salvage hypoxanthine and guanine to make IMP and GMP, PRPP accumulation, activation of Amido phosphoribosyltransferase and excessive purine denovo synthesis. Lesch-Nyhan syndrome is characterized by gout( joint pain) , kidney stones , severe self-mutilation, spastic movements, extreme hostility & mental retardation.  Digestion & Degradation of nucleic acid/Purine Nucleotides in G.I.T Ingested nucleic acids are degraded in G.I.T. to nucleotides by pancreatic nucleases & intestinal phosphodiesterases Nucleotides are then converted to nucleosides by base-specific nucleotidases & non-specific phosphatases Then nucleoside are directly absorbed & further degraded by nucleosidases or nucleoside phosphorylases to release the purine base as follow: Nucleoside(Guanosine)+H2O→base(Guanine)+ ribose (catalysed by nucleosidase) Nucleoside(e.g Guanosine, Inosine) + Pi→ base + ribose-1-phosphate (nucleoside phosphorylase) Ribose 1-phosphate is converted to ribose 5-phosphate (used in PRPP synthesis)by isomerase. some bases are reused to form nucleotides by salvage pathways Most bases are degraded to Uric acid & are excreted in urine in human being. Digestion & Degradation of nucleic acid/Purine Nucleotides in G.I.T Purines in humans are degraded to Urate  AMP Degradation/Catabolism AMP is degraded to IMP by AMP deaminase Nucleotidase also catalize (hydrolytic cleavage of the glycosidic bond) breakdown AMP to Adenosine & IMP to Inosine Adenosine is deaminated to inosine by adenosine deaminase. Inosine are converted by purine nucleoside phosphorylase to hypoxanthine & ribose 1- phosphate. Xanthine oxidase oxidizes hypoxanthine to xanthine & then xanthine to uric acid excreted in urine in human. End product of purine metabolism in humans is uric acid But uric acid can lose a proton at physiological pH to form insoluble urate.  GMP degradation/Catabolism Nucleotidase catalyze breakdown of GMP to Guanosine Nucleoside Phosphorylase catalyze breakdown of Guanosine to Guanine & Ribose-1-phosphate Guanase:Deaminates Guanine to form xanthine Xanthine Oxidase: oxidizes xanthine to uric acid,excreted in the urine Xanthine Oxidase enzyme contains FAD, Molybdenum & Iron, & is mainly found in liver & small intestine.  The Fate of Uric Acid In primates(human), dogs, birds, reptiles metabolism stops in uric acid and excreted In mollusc and in mammals that are not primates, it is oxidized by urate oxidase to allantoin and excreted. In bony fishes (teleosts), allantoin is hydrolyzed by allantoinase to allantoic acid and excreted Cartilaginous fish (sharks and rays) and amphibians it is stops as glyoxylic acid and two equivalents of urea catalyzed by allantoicase. Degradation of Uric Acid Excreted or degraded to various levels depending on the species – Marine invertebrates can break down uric acid all the way down to ammonia Organisms that do not excrete urea can remove excess nitrogen through uric acid – Complicated reactions but conserved water (uric acid barely soluble)  Pyrimidine Metabolism Pyrimidine biosynthesis – De novo pathways – Salvage pathway Pyrimidine catabolism  De novo synthesis of Pyrimidine nucleotide Types of Pyrimidine Bases: Cytosine, Thymine, Uracil. De novo synthesis of pyrimidines: Pyrimidine ring is synthesized first before ribose-5-phosphate is attached to form a pyrimidine nucleotide in the cytosol. Sources of different atoms of Pyrimidine Ring BCH 212  De novo biosynthesis of pyrimidines Step 1 : Synthesis of carbamoyl phosphate from bicarbonate , ammonia & hydrolysis of 2 ATP ,a reaction catalyzed by carbamoyl phosphate synthetase (CPS)II. Occurs in 3 stages: Bicarbonate is phosphorylated by ATP to form carboxyphosphate & ADP. Ammonia (usually produced from hydrolysis of the side chain of glutamine) then reacts with carboxyphosphate to form carbamic acid & inorganic phosphate. carbamic acid is phosphorylated by another ATP to form carbamoyl phosphate. CPS II in bacteria has 3 active sites & intermediates generated at one site move to the next without leaving the enzyme(substrate channeling),to prevents loss of intermediates by diffusion & prevent hydrolysis of labile intermediates such as carboxyphosphate & carbamic acid (which decomposes in less than 1 sec at pH 7). Step 2: Carbamoyl phosphate reacts with Aspartate to form N- carbamoyl aspartate. Catalyzed by Aspartate Transcarbamoylase (ATCase). Is the committed step in pyrimidine biosynthesis BCH 212  De novo biosynthesis of pyrimidines-Cont’d Step 3: Dihydroorotase enzyme act on N-carbamoyl aspartate to remove water & form dihydroorotate Note: In mammals, the 1St three enzymes :Carbamoyl phosphate synthetase II, Aspartate transcarbamoylase, & Dihydroorotase— are part of a single trifunctional CAD protein , which contains three identical polypeptide chains with active sites for all three reactions. Step 4: Dihydroorotate is then dehydrogenated(oxidized) by Coenzyme quinone (NAD+) to form Orotate (pyrimidine ring) in a reaction catalyzed by dihydroorotate dehydrogenase enzyme. Plasmodium falciparum Dihydroorotate dehydrogenase (PfDHODH) is a target for novel antimalarial drug development  De novo biosynthesis of pyrimidines-Cont’d Step 5: Orotate reacts with 5- phosphoribosyl-1- pyrophosphate(PRPP) to form orotidylate/orotidine 5’- monophosphate(OMP) Catalyzed by orotate phosphoribosyl- transferase enzyme PRPP provides the ribose 5-phosphate side chain. Step 6: Orotidylate(OMP) is decarboxylated to form uridylate/Uridine monophosphate (UMP),catalyzed by the enzyme orotidylate decarboxylase. orotate-phosphoribosyltransferase & decarboxylase form UMP synthase. Overview of the De novo biosynthesis of Pyrimidine  De novo biosynthesis of pyrimidines-Cont’d Step 7: Uridylate (UMP) is converted to uridine 5'- triphosphate (UTP) in 2 stages using 2 ATP and kinases enzyme(specific UMP kinase & less specific UDP kinase) Pyrimidine nucleoside diphosphates & triphosphates are interconverted by less/broad specific nucleoside diphosphate kinase(e.g UDP kinase), it also catalyze the interconversion of several ribonucleosides or even deoxyribonucleosides.  Synthesis of Cytidine Triphosphate(CTP) from UTP CTP is formed from UTP by replacing a carbonyl with amino group on UTP via the action of CTP synthetase using a ATP. In animals nitrogen donor is glutamine. In bacteria, nitrogen donor is ammonia. Overview of Biosynthesis of thymidylate (dTMP) Source: Lehninger Principles Of Biochemistry 4th Edition  Salvage Pathway synthesis of Pyrimidine Free pyrimidine released in cells during the metabolic breakdown of nucleotides are salvaged and recycled to make pyrimidine nucleotides. Salvage synthesis of pyrimidine is catalyzed by pyrimidine phosphoribosyl transferase enzyme. pyrimidine phosphoribosyl transferase uses 5-phosphoribosyl-1- pyrophosphate(PRPP) as the source of ribose-5-phosphate.  Feedback regulatory control of Pyrimidine Synthesis In Bacteria regulation occur at aspartate transcarbamoylase(ATCase),that is feedback inhibited by CTP but activated by ATP. In Animals – regulation is at carbamoyl phosphate synthetase II(CPS II) – UDP & UTP inhibit CPS II - ATP & PRPP activate CPS II. OMP decarboxylase is competitively inhibited by UMP & CMP, also controls pyrimidine formation Inhibition of PRPP synthetase by ADP & GDP in purine synthesis regulation, also regulates pyrimidine synthesis Overview of regulation of pyrimidine Biosysnthesis  Catabolism/degradation of pyrimidine nucleotide Pyrimidine nucleotides (CMP & UMP) are dephosphorylated to form nucleoside by nucleotidase. nucleosides are cleaved to produce ribose 1- phosphate & free pyrimidine bases(cytosine, uracil, & thymine) by nucleosidase & phosporylase. Cytosine is deaminated by deaminase to form uracil, which is degraded to CO2,NH4, β-alanine. Thymine is degraded to CO2,NH4 & β-aminoisobutyrate These products of pyrimidine degradation are excreted in the urine or converted to CO2, H2O, and NH4+ (which forms urea).  Some clinical inhibitors of nucleotide synthesis 6-Mercaptopurine(anticancer drug) inhibits the conversion of IMP to AMP & GMP. Also feed back inhibits glutamine PRPP amidotransferase. Sulfonamides(antibiotics) :analogs of para aminobenzoic acid (PABA) that inhibits synthesis of folic acid(THF) & reactions of purine synthesis requiring folic acid by inhibiting GAR transformylase & AICAR transformylase) in bacteria. This inhibition causes accumulation of 5-aminoimidazole-4- carboxamide ribonucleotide & is reversed by the addition of para-aminobenzoate. Acyclovir (Anti-viral drugs for chickenpox & STD/herpes): Is converted to triphosphate form that inhibits viral DNA polymerase responsible for DNA/RNA polymerization(synthesis). Methotrexate (anticancer drug) :Is an analogue of folic acid that inhibits dihydrofolate reductase & thymidylate synthase to prevent tetrahydrofoliate formation & inhibit the reaction requiring folic acid in purine denovo synthesis & thymidine synthesis. Cancer cells requires more dTMP for DNA replication, inhibiting thymidylate synthase decrease dTMP production & prevent growth of cancer. Folinic acid (Leucovorin) is given along with methotrexate to for normal cells to get required thymidine nucleotides.  Some clinical inhibitors of nucleotide synthesis Azaserine is a glutamine antagonist & inhibits reactions in purine & pyrimidine synthesis using glutamine as substrate. Used as a potential antineoplastic/anti-tumor agent. Mycophenolic acid: Is a reversible non-competative inhibitor of IMP dehydrogenase(enzyme converting IMP to XMP). It deprive rapidly proliferating T and B lymphocyte cells of guanine nucleotides. Used as immunosuppressant drug to prevent organ rejection in organ transplantation 6-Thioguanine & 8 aza guanine (anticancer agents):inhibits glutamine PRPP amidotransferase a n d also inhibit conversion of IMP to GMP. Fluorouracil (anticancer drug): is converted to Fluorodeoxyuridine monophosphate (FdUMP) ,which irreversibly inhibits thymidylate synthase(enzyme that converts dUMP to dTMP). Zidovudine (ZDV), formerly called AZT: HIV/AIDs and analogue of deoxythymidine in DNA used to terminate nucleotides formation by inhibiting the viral DNA polymerase in herpes virus & AIDS virus  Abnormalities in Nucleic acid Metabolism 1. Gout: Painful arthritis mostly in male caused by increased formation of uric acid in the blood or decreased renal excretion resulting in urate deposit at joints/under skin. Urate crystals also appear as kidney stones Purine-rich foods (shellfish, eggs rich in nucleic acids) may exacerbate the condition. TREATMENT : Allopurinol, a competitive inhibitor of xanthine oxidase(enzyme that degrades purines to uric acid). Allopurinol (a structural analog of hypoxanthine) is a substrate for xanthine oxidase. It is converted to oxypurinol (alloxanthine), which remains tightly bound to xanthine oxidase, preventing further catalytic activity. When xanthine oxidase is inhibited, uric acid is not formed, the excreted products of purine metabolism are xanthine & hypoxanthine,which are more soluble In patients with normal levels of HGPRT, hypoxanthine can be salvaged, reducing PRPP levels & de-novo purine synthesis. Hypoxanthine & xanthine do not accumulate to harmful concentrations because they are more soluble, less likely to initiate an inflammatory response , more easily excreted & do not form urate deposits Allopurinol is sometimes given to leukemia & Iymphoma patients along anti-cancer drugs as Adjunct therapy to enhance retention and potentiation of anti-cancer drugs ,since cancer drugs may be deactivated by xanthine oxidase, allopurinol also inhibit xanthine oxidase to prevent urate high level caused by increased death of cancer cells and breakdown of their nucleic acids due to cancer therapies. Allopurinol also prevents the formation of kidney stones and blocks other deleterious consequences of hyperuricemia by preventing the formation of urate Colchicine(anti-inflammatory) & uricosuric(increases renal excretion of uric acid) drugs also remove urates from the joint & body tissues Gout  Abnormalities in Nucleic acid metabolism- Cont’d 2. Lesch-Nyhan syndrome. Genetic disorder in male(X-linked recessive), characterized by cognitive deficits, extremely aggressive behaviour & self- mutilation caused by deficiency of hypoxanthine-guanine phosphoribosyltransferase(HGPRT), leads to inability to salvage hypoxanthine or guanine, high PRPP level, increased purine de-novo synthesis. Excess purines are broken down into uric acid & over production of uric acid/urate (hyperuricemia) & kidney stones. Treatment(relieve the self-injurious behaviors): Behavioural therapy , intraoral protective appliances(lip shields), dental extraction, protective strapes/helmets ,drugs like Ecopipam(D1 Dopamine Receptor Antagonist) & Aminoimidazole Carboxamide Riboside (AICAR),a (Precursor of Purine ) are at various stages of clinical trials. BCH 212 59  Abnormalities in Nucleic acid metabolism- Cont’d 3. Von Gierke’s(Type 1 glycogen-storage) hereditary disease: Due to deficiency of glucose-6-phosphatase(converts a glycogen intermediate glucose-6-phosphate to glucose in liver). Leads to hypoglycemia, liver tumor, increased production of ribose 5- phosphate(PRPP precursor ),purine overproduction, lactic acidosis & hyperuricemia (elevated urate level). Treatment: frequent feedings of glucose or cornstarch & allopurinol to prevent uric acid deposition in kidneys and joints.  Abnormalities in Nucleic acid metabolism- Cont’d 5. xanthine oxidase deficiency : Associated with Hypouricemia & increased excretion of hypoxanthine and xanthine. It is caused to a genetic defect or severe liver damage. Treatment: Enzyme replacement therapy AMP Degradation/Catabolism Abnormalities in Nucleic acid metabolism- Cont’d dysfunction pyrimidine metabolism OROTIC ACIDURIA: inherited disorder caused by deficiency of either or both orotate-phosphoribosyl transferase and decarboxylase(catalytic domains of uridine Monophosphate synthase, that convert orotate to OMP & OMP to UMP),leading to excessive excretion of orotate in urine & megaloblastic anemia(anemia associated with vitamin B12 or folate deficiency). Treatment: life time usage of uridine (pyrimidine-analog containing uracil) that bypass the missing enzymes & can be salvaged to UMP, which can be converted to all the other pyrimidines.  Some clinical relevance of nucleotide synthesis Thymidylate synthesis is crucial in cell proliferation. Hence, the enzymes involved in this biosynthetic pathway are targets for chemotherapeutic and antibiotic drug design. Chemotherapeutic agents such as 5-fluorouracil (5-F-dUMP) and Raltitrexed target classical TSase for skin, colon, and ovarian cancers. Methotrexate, used clinically for treating cancer and autoimmune disorders, targets DHFR and trimethoprim is specific to bacterial DHFR inhibition. Until 2002 thymidylate was thought to be synthesized either de novo by TSase or by scavenging exogenous thymidine compounds by the action of thymidine kinase (Tdk). Genomic analysis on archaea and bacteria revealed many organisms lacking thyA do not carry folA as well as the gene coding for thymidine kinase, yet they survived in thymidine deficient media. This observation led to the discovery of a new gene thyX, which codes for a different class of thymidylate synthases: Flavin-Dependent Thymidylate Synthase (FDTS, EC 2.1.1.148).