Installation of the pyrrolyl-2-carboxyl pharmacophore by CouN1 and CouN7 in the late biosynthetic steps of the aminocoumarin antibiotics clorobiocin and coumermycin A1.
The 5-methyl-2-pyrrolylcarbonyl moiety of the aminocoumarin antibiotics clorobiocin and coumermycin A1 is the key pharmacophore for targeting the ATP-binding site of GyrB for inhibition of the bacterial type-II topoisomerase DNA gyrase. During the late stage of clorobiocin and coumermycin A1 biosynthesis, the pyrrolyl-2-carboxyl group is transferred from the peptidyl carrier proteins Clo/CouN1 to the 3'-hydroxyl of the 4-methoxy-L-noviosyl scaffold by the action of the acyltransferases Clo/CouN7. CouN1 and CouN7 have now been heterologously expressed and purified from Escherichia coli. The apo form of CouN1 is converted to the acyl-holo form by loading with pyrrolyl-2-carboxyl-S-pantetheinyl moieties from synthetic pyrrolyl- and 5-methylpyrrolyl-CoAs by the action of the phosphopantetheinyl transferase Sfp. CouN7 acts as an acyltransferase, moving the pyrrole acyl moieties from CouN1 to the noviose sugar of descarbamoylnovobiocin. When the 5-methylpyrrolyl-2-carboxyl-thioester of CouN1 is the cosubstrate, the in vitro product differs from clorobiocin only in a CH3 for Cl group change on the coumarin ring. Double transfer of this acyl moiety by CouN7 to the penultimate intermediate in coumermycin A1 assembly completes that antibiotic biosynthetic pathway.
Garneau-Tsodikova
Sylvie
S
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA.
Stapon
Anthony
A
Kahne
Daniel
D
Walsh
Christopher T
CT
D000217:Acyltransferases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D000249:Adenosine Monophosphate / Q000378:metabolism; D049933:Aminocoumarins / Q000378:metabolism*; D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis*; D002384:Catalysis; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005800:Genes, Fungal; D007700:Kinetics; D009675:Novobiocin / Q000031:analogs & derivatives* / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics
D049933:Aminocoumarins; D000903:Antibiotics, Antineoplastic; D011994:Recombinant Proteins; D009675:Novobiocin; C006260:clorobiocin; D000249:Adenosine Monophosphate; D000217:Acyltransferases; C004628:coumermycin