Novobiocin, also known asalbamycin, is anaminocoumarin antibiotic that is produced by the actinomyceteStreptomyces niveus, which has recently been identified as a subjective synonym forS. spheroides[1] a member of the classActinomycetia. Other aminocoumarin antibiotics include clorobiocin and coumermycin A1.[2] Novobiocin was first reported in the mid-1950s (then calledstreptonivicin).[3][4]
Novobiocin was licensed for clinical use under the tradename Albamycin (Upjohn) in the 1960s. Itsefficacy has been demonstrated inpreclinical andclinical trials.[5][6] The oral form of the drug has since been withdrawn from the market due to lack of efficacy.[7] A combination product of novobiocin and tetracycline, sold by Upjohn under brand names such as Panalba and Albamycin-T, was in particular the subject of intense FDA scrutiny before it was finally taken off the market.[8][9] Novobiocin is an effectiveantistaphylococcal agent used in the treatment ofMRSA.[10]
The molecular basis of action ofnovobiocin, and other related drugsclorobiocin andcoumermycin A1 has been examined.[2][11][12][13][14] Aminocoumarins are verypotent inhibitors of bacterial DNA gyrase and work by targeting the GyrB subunit of the enzyme involved in energy transduction. Novobiocin as well as the otheraminocoumarinantibiotics act as competitive inhibitors of theATPase reaction catalysed by GyrB. The potency of novobiocin is considerably higher than that of thefluoroquinolones that also targetDNA gyrase, but at a different site on the enzyme. The GyrA subunit is involved in the DNA nicking and ligation activity.[citation needed]
Novobiocin has been shown to weakly inhibit the C-terminus of the eukaryotic Hsp90 protein (high micromolar IC50). Modification of the novobiocin scaffold has led to more selective Hsp90 inhibitors.[15] Novobiocin has also been shown to bind and activate the Gram-negative lipopolysaccharide transporter LptBFGC.[16][17]
The ATP binding pocket of polymerase theta is blocked by novobiocin resulting in a loss of ATPase activity. This results in the loss of microhomology-mediated end joining as a pathway for homologous recombination deficient cells to circumvent DNA damaging agents. The action of novobiocin is syngeristic with PARP inhibitors for reducing tumor size in a mouse model.[18]
Novobiocin is an aminocoumarin. Novobiocin may be divided up into three entities; a benzoic acid derivative, a coumarin residue, and the sugar novobiose.[11]X-ray crystallographic studies have found that the drug-receptor complex of Novobiocin and DNA Gyrase shows that ATP and Novobiocin have overlapping binding sites on the gyrase molecule.[19] The overlap of the coumarin and ATP-binding sites is consistent with aminocoumarins being competitive inhibitors of the ATPase activity.[20]
Thisaminocoumarin antibiotic consists of three major substituents. The 3-dimethylallyl-4-hydroxybenzoic acid moiety, known as ring A, is derived fromprephenate anddimethylallyl pyrophosphate. The aminocoumarin moiety, known as ring B, is derived fromL-tyrosine. The final component of novobiocin is the sugar derivativeL-noviose, known as ring C, which is derived from glucose-1-phosphate. The biosynthetic gene cluster for novobiocin was identified by Heide and coworkers in 1999 (published 2000) fromStreptomyces spheroides NCIB 11891.[21] They identified 23 putative open reading frames (ORFs) and more than 11 other ORFs that may play a role in novobiocin biosynthesis.[citation needed]
The biosynthesis of ring A (seeFig. 1) begins with prephenate which is a derived from theshikimic acid biosynthetic pathway. The enzyme NovF catalyzes the decarboxylation ofprephenate while simultaneously reducing nicotinamide adenine dinucleotide phosphate (NADP+) to produceNADPH. Following this NovQ catalyzes the electrophilic substitution of the phenyl ring withdimethylallyl pyrophosphate (DMAPP) otherwise known as prenylation.[22] DMAPP can come from either the mevalonic acid pathway or the deoxyxylulose biosynthetic pathway. Next the 3-dimethylallyl-4-hydroxybenzoate molecule is subjected to two oxidative decarboxylations by NovR and molecular oxygen.[23] NovR is a non-heme iron oxygenase with a unique bifunctional catalysis. In the first stage both oxygens are incorporated from the molecular oxygen while in the second step only one is incorporated as determined by isotope labeling studies. This completes the formation of ring A.
Figure 1. Biosynthetic scheme of benzamide portion of novobiocin (4-hydroxy-3-(3-methylbut-2-en-1-yl)benzoic acid)
The biosynthesis of ring B (seeFig. 2) begins with the natural amino acidL-tyrosine. This is then adenylated and thioesterified onto the peptidyl carrier protein (PCP) of NovH byATP and NovH itself.[24] NovI then further modifies this PCP bound molecule by oxidizing the β-position usingNADPH and molecular oxygen. NovJ and NovK form a heterodimer of J2K2 which is the active form of this benzylic oxygenase.[25] This process uses NADP+ as a hydride acceptor in the oxidation of the β-alcohol. This ketone will prefer to exist in its enol tautomer in solution. Next a still unidentified protein catalyzes the selective oxidation of the benzene (as shown in Fig. 2). Upon oxidation this intermediate will spontaneously lactonize to form the aromatic ring B and lose NovH in the process.
Figure 2. Biosynthesis of 3-amino-4,7-dihydroxy-2H-chromen-2-one component of novobiocin (ringB)
The biosynthesis ofL-noviose (ring C) is shown inFig. 3. This process starts from glucose-1-phosphate where NovV takes dTTP and replaces the phosphate group with a dTDP group. NovT then oxidizes the 4-hydroxy group using NAD+. NovT also accomplishes a dehydroxylation of the 6 position of the sugar. NovW then epimerizes the 3 position of the sugar.[26] The methylation of the 5 position is accomplished by NovU andS-adenosyl methionine (SAM). Finally NovS reduces the 4 position again to achieve epimerization of that position from the starting glucose-1-phosphate usingNADH.
Figure 3. Biosynthesis of L-noviose component of novobiocin (ringC)
Rings A, B, and C are coupled together and modified to give the finished novobiocin molecule. Rings A and B are coupled together by the enzyme NovL usingATP to diphosphorylate the carboxylate group of ring A so that the carbonyl can be attacked by the amine group on ring B. The resulting compound is methylated by NovO and SAM prior to glycosylation.[27] NovM adds ring C (L-noviose) to the hydroxyl group derived from tyrosine with the loss of dTDP. Another methylation is accomplished by NovP and SAM at the 4 position of theL-noviose sugar.[28] This methylation allows NovN to carbamylate the 3 position of the sugar as shown inFig. 4 completing the biosynthesis of novobiocin.
Figure 4. Completed biosynthesis of novobiocin from ring systemsA,B, andC.
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^Raad II, Hachem RY, Abi-Said D, Rolston KV, Whimbey E, Buzaid AC, Legha S (January 1998). "A prospective crossover randomized trial of novobiocin and rifampin prophylaxis for the prevention of intravascular catheter infections in cancer patients treated with interleukin-2".Cancer.82 (2):403–11.doi:10.1002/(SICI)1097-0142(19980115)82:2<412::AID-CNCR22>3.0.CO;2-0.PMID9445199.S2CID10940970.
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^Yu XM, Shen G, Neckers L, Blake H, Holzbeierlein J, Cronk B, Blagg BS (September 2005). "Hsp90 inhibitors identified from a library of novobiocin analogues".Journal of the American Chemical Society.127 (37):12778–9.Bibcode:2005JAChS.12712778Y.doi:10.1021/ja0535864.PMID16159253.
^Tsai FT, Singh OM, Skarzynski T, Wonacott AJ, Weston S, Tucker A, Pauptit RA, Breeze AL, Poyser JP, O'Brien R, Ladbury JE, Wigley DB (May 1997). "The high-resolution crystal structure of a 24-kDa gyrase B fragment from E. coli complexed with one of the most potent coumarin inhibitors, clorobiocin".Proteins.28 (1):41–52.doi:10.1002/(sici)1097-0134(199705)28:1<41::aid-prot4>3.3.co;2-b.PMID9144789.
^Pacholec M, Hillson NJ, Walsh CT (September 2005). "NovJ/NovK catalyze benzylic oxidation of a beta-hydroxyl tyrosyl-S-pantetheinyl enzyme during aminocoumarin ring formation in novobiocin biosynthesis".Biochemistry.44 (38):12819–26.CiteSeerX10.1.1.569.1481.doi:10.1021/bi051297m.PMID16171397.
^Thuy TT, Lee HC, Kim CG, Heide L, Sohng JK (April 2005). "Functional characterizations of novWUS involved in novobiocin biosynthesis from Streptomyces spheroides".Archives of Biochemistry and Biophysics.436 (1):161–7.doi:10.1016/j.abb.2005.01.012.PMID15752721.
^Pacholec M, Tao J, Walsh CT (November 2005). "CouO and NovO: C-methyltransferases for tailoring the aminocoumarin scaffold in coumermycin and novobiocin antibiotic biosynthesis".Biochemistry.44 (45):14969–76.doi:10.1021/bi051599o.PMID16274243.
^Freel Meyers CL, Oberthür M, Xu H, Heide L, Kahne D, Walsh CT (January 2004). "Characterization of NovP and NovN: completion of novobiocin biosynthesis by sequential tailoring of the noviosyl ring".Angewandte Chemie.43 (1):67–70.doi:10.1002/anie.200352626.PMID14694473.
