Tunicamycin homologues have varying molecular weights owing to the variability in fatty acid side chain conjugates.[3]
Biosynthesis
The biosynthesis of tunicamycins was studied in Streptomyces chartreusis and a proposed biosynthetic pathway was characterized. The bacteria utilize the enzymes in the tun gene cluster (TunA-N) to make tunicamycins.[4]
TunA uses the starter unit uridine diphosphate-N-acetyl-glucosamine (UDP-GlcNAc) and catalyzes the dehydration of the 6’ hydroxyl group. First, a Tyr residue in TunA abstracts a proton from the 4’ hydroxyl group, forming a ketone at that position. A hydride is subsequently abstracted from the 4’ carbon by NAD+, forming NADH. The ketone is stabilized by hydrogen bonding from the Tyr residue, and a nearby Thr residue. A glutamate residue then abstracts a proton from the 5’ carbon, pushing the electrons up to form a double bond between the 5’ and 6’ carbon. A nearby cysteine donates a proton to the hydroxyl group as it leaves as water. NADH donates a hydride to the 4’ carbon, reforming a hydroxide in that position and forming UDP-6’-deoxy-5-6-ene-GlcNAc. TunF then catalyzes the epimerization of the intermediate to UDP-6’-deoxy-5-6-ene-GalNAc, changing the 4’ hydroxyl from the equatorial to axial position.[5]
The other starter unit for tunicamycin is uridine, which is produced from uridine triphosphate (UTP). TunN is a nucleotide diphosphatase, and catalyzes the removal of pyrophosphate from UTP to form uridine monophosphate. The last phosphate is removed by the putative monophosphatase, TunG.
Once uridine and UDP-6’-deoxy-5-6-ene-GalNAc are produced, TunB catalyzes their linkage at the 6’ carbon of UDP-6’-deoxy-5-6-ene-GalNAc. TunB uses S-adenyslmethionine (SAM) to form a radical on the 5’ carbon of the ribose on uracil. TunM is thought to catalyze the formation of a new bond between the 5’ carbon of uridine and the 6’ carbon of UDP-6’-deoxy-5-6-ene-GalNAc using the electron from the uridine radical and one of the electrons from the double bond of UDP-6’-deoxy-5-6-ene-GalNAc. The radical on UDP-6’-deoxy-5-6-ene-GalNAc is then quenched by abstracting a hydrogen from SAM.[6] The resulting molecule is UDP-N-acetyl-tunicamine. TunH then catalyzes the hydrolysis of UDP from UDP-N-acetyl-tunicamine. Another molecule of UDP-GlcNAc is introduced, and a β-1,1 glycosidic bond is subsequently formed, catalyzed by TunD. The resulting molecule is deacetylated by TunE. TunL and a fatty acyl-ACP ligase are used to load metabolic fatty acids onto the acyl carrier protein, TunK. TunC then attaches the fatty acid to the free amine, producing tunicamycin.
See also
Glycosylation - tunicamycin blocks all N-glycosylation of proteins
^Wyszynski F, Hesketh A, Bibb M, Davis B (2010). "Dissecting tunicamycin biosynthesis by genome mining: cloning and heterologous expression of a minimal gene cluster". Chemical Science. 1 (5): 581. doi:10.1039/c0sc00325e.
^Wyszynski F, Lee S, Yabe T, Wang H, Gomez-Escribano JP, Bibb M (July 2012). "Biosynthesis of the tunicamycin antibiotics proceeds via unique exo-glycal intermediates". Nature Chemistry. 4 (7): 539–546. Bibcode:2012NatCh...4..539W. doi:10.1038/nchem.1351. PMID22717438.
^Giese B (August 1989). "The Stereoselectivity of Intermolecular Free Radical Reactions [New Synthetic Methods (78)]". Angewandte Chemie International Edition in English. 28 (8): 969–980. doi:10.1002/anie.198909693.