Regiospecific chlorination of (S)-beta-tyrosyl-S-carrier protein catalyzed by SgcC3 in the biosynthesis of the enediyne antitumor antibiotic C-1027.
C-1027 is a potent antitumor antibiotic composed of an apo-protein and a reactive enediyne chromophore. The chromophore consists of four different chemical subunits including an (S)-3-chloro-4,5-dihydroxy-beta-phenylalanine moiety, the biosynthesis of which from l-alpha-tyrosine is catalyzed by six proteins, SgcC, SgcC1, SgcC2, SgcC3, SgcC4, and SgcC5. Biochemical characterization of SgcC3 unveiled the following: (i) SgcC3 is a flavin adenine dinucleotide (FAD)-dependent halogenase; (ii) SgcC3 acts only on the SgcC2 peptidyl carrier protein-tethered substrates; (iii) SgcC3-catalyzed halogenation requires O2 and reduced FAD and either the C-1027 pathway-specific flavin reductase SgcE6 or E. coli flavin reductase (Fre) can support the SgcC3 activity; (iv) SgcC3 also efficiently catalyzes bromination but not fluorination or iodination; (v) SgcC3 can utilize both (S)- and (R)-beta-tyrosyl-S-SgcC2 but not 3-hydroxy-beta-tyrosyl-S-SgcC2 as a substrate. These results establish that SgcC3 catalyzes the third enzymatic transformation during the biosynthesis of the (S)-3-chloro-4,5-dihydroxy-beta-phenylalanine moiety of C-1027 from l-alpha-tyrosine. SgcC3 now represents the second biochemically characterized flavin-dependent halogenase that acts on a carrier protein-tethered substrate. These findings will facilitate the engineering of new C-1027 analogs by combinatorial biosynthesis methods.
Lin
Shuangjun
S
Division of Pharmaceutical Sciences, University of Wisconsin National Cooperative Drug Discovery Group, and Department of Chemistry, University of Wisconsin-Madison, Madison, Wisconsin 53705, USA.
Van Lanen
Steven G
SG
Shen
Ben
B
D000617:Aminoglycosides / Q000737:chemistry*; D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D002352:Carrier Proteins / Q000378:metabolism*; D002384:Catalysis; D002713:Chlorine / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D053281:Enediynes / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D005415:Flavins / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D015394:Molecular Structure; D010649:Phenylalanine / Q000737:chemistry / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism
D000617:Aminoglycosides; D000903:Antibiotics, Antineoplastic; D002352:Carrier Proteins; D053281:Enediynes; D005415:Flavins; C058024:C 1027; D014443:Tyrosine; D010649:Phenylalanine; D002713:Chlorine