Back Cover: Insights into the Biosynthesis of Dehydroalanines in Goadsporin (ChemBioChem 3/2016)

Abstract

The back cover picture shows a paper disk diffusion assay of goadsporin against Streptomyces lividans TK23 on an agar plate. Through the inactivation of godF and godG, goadsporin B and goadsporin C were identified. The structures of these compounds provide insights into the mechanism of dehydroamino acid formation catalyzed by LanB‐type enzymes. More information can be found in the communication by H. Onaka et al. on page 218 in Issue 3, 2016 (DOI: 10.1002/cbic.201500541).

YcaO-Dependent Posttranslational Amide Activation: Biosynthesis, Structure, and Function

“…The structure of goadsporin is unusual for a LAP because in addition to its azole heterocycles it contains two dehydrated amino acids (Figure ). , Genetic deletion studies support a biosynthetic pathway in which the azoles are formed first by a fused cyclodehydratase (GodD) and a dehydrogenase (GodE) . In the next likely step, dehydroamino acids are formed by what is now termed a split LanB dehydratase that catalyzes the glutamylation of select Ser and Thr (GodG) and subsequent elimination (GodF) to yield the corresponding dehydroalanine and dehydrobutyrine moieties. , Putative membrane-associated proteases (GodB and GodC) cleave the modified precursor peptide (GodA), and an N-acetyltransferase (GodH) modifies the newly formed N-terminus. , Transcription of the goadsporin BGC is controlled by GodR, an activator, whose overexpression results in increased production of goadsporin, although the best titers were achieved through heterologous expression in S. lividans and coculturing with Tsukamurella pulmonis . , In order to prevent inhibition of its own growth, the BGC encodes the immunity protein GodI (see below).…”

“…Thus, a LanB dehydratase can be found in multiple RiPP classes and can provide increased chemical/structural diversity to a natural product pathway. Indeed, the dehydroamino acids of goadsporin are required for bioactivity . Other structural features that have been investigated show flexibility in swapping oxazole and methyloxazole heterocycles, but alteration of the C-terminus or to Gly10 compromise its activity.…”

“…The roles of the aforementioned core biosynthetic enzymes were first assigned through bioinformatic analysis and characterization of partially processed intermediates in genetically manipulated strains. ,,,, Although not conclusive, these studies suggested the biosynthetic order of events began with azol­(in)­e formation, then Ser/Thr dehydration, followed by macrocyclization through a putative [4 + 2] cycloaddition. The cyclodehydratase (G) was readily identified based on its obvious homology to azoline-forming YcaO proteins, , and similarly, the role of the dehydrogenase (E) in azoline oxidation to their corresponding azole heterocycles was easily recognized via homology with other flavin-dependent dehydrogenases known to dehydrogenate azoline heterocycles in RiPP pathways (Figure ).…”

“…These proteins catalyze dehydration through glutamylation of the hydroxyl group (B protein) using glutamyl-tRNA Glu with subsequent elimination (C protein). ,, A formal [4 + 2] cycloaddition of two dehydroalanines forms the six-membered nitrogenous heterocycle (Figure ). , The identity of the [4 + 2] cycloaddition enzyme was inferred by its absence in goadsporin (section ), , which structurally resembles a presumed intermediate during thiopeptide biosynthesis. Deleting this gene in thiopeptide BGCs resulted in the isolation of linear intermediates lacking this 6-membered heterocycle. ,, Unexpectedly, the [4 + 2] enzyme has high similarity to the elimination domain of LanB dehydratases and possibly evolved through a duplication event.…”

“…For example, in cyanobactin pathways, when a prenyltransferase is present, certain Ser/Thr residues must remain uncyclized in order for it to be able to act on the substrate and may explain why TruD is a Cys-selective cyclodehydratase since tru pathways tend to include prenylation . Similar selectivity is demonstrated by the azole synthetase (GodD/E) from goadsporin biosynthesis (section ) which processes all Cys and Thr but only one of four Ser are converted to oxazoles in the mature compound. , Of the other three Ser residues, two become dehydroalanine while the third remains unmodified. , As for thiopeptide biosynthetic pathways (section ), the cyclodehydratase skips over Cys involved in side ring macrocycles (e.g., nosiheptide or lactocillin), but the basis for this site-specificity has not been explored. More investigated is the regioselectivity during Ser/Thr cyclodehydration.…”