A Remarkable Oxidative Cascade That Replaces the Riboflavin C8 Methyl with an Amino Group during Roseoflavin Biosynthesis

Abstract

Roseoflavin is a naturally occurring riboflavin analogue with antibiotic properties. It is biosynthesized from riboflavin in a reaction involving replacement of the C8 methyl with a dimethylamino group. Herein we report the identification of a flavin-dependent enzyme that converts flavin mononucleotide (FMN) and glutamate to 8-amino-FMN via the intermediacy of 8-formyl-FMN. A mechanistic proposal for this remarkable transformation is proposed.

The Crystal Structure of RosB: Insights into the Reaction Mechanism of the First Member of a Family of Flavodoxin‐like Enzymes

“…(Figure 5a nd Figure S7). [10] Aw ater molecule close to C8 is appropriately positioned to convert OHC-RP into HO 2 C-RP.Furthermore, structural and mutagenesis data allowed to plausibly model glutamate in front of C8 of RP providing as tructural frame for the transamination reaction ( Figure S8). This (partially) deprotonated H 2 O abstracts the C8 methyl proton which formally leads to ar educed flavin.…”

“…O 2 reacts with RP and the resulting C8 methyl hydroperoxide anion is protonated and dissociates to OHC-RP and OH À being in line with the fact that H 2 O 2 was neither detected by us [9] nor by others. [10] Aw ater molecule close to C8 is appropriately positioned to convert OHC-RP into HO 2 C-RP.Furthermore, structural and mutagenesis data allowed to plausibly model glutamate in front of C8 of RP providing as tructural frame for the transamination reaction ( Figure S8).…”

“…[9] Thes ubstrate for RosB was found to be RP (not riboflavin) and the final reaction product of RosB was found to be AFP (not AF). Another research team also reported on the identification of RosB as the key enzyme of roseoflavin biosynthesis, [10] ar equirement of thiamine,however,w as not mentioned. Depending on the assay conditions different stable reaction intermediates are formed by RosB (Figure 1b).…”

Die Kristallstruktur von RosB: Einblicke in den Reaktionsmechanismus des ersten Mitglieds einer flavodoxinähnlichen Enzymfamilie

“…Auf die Decarboxylierung von HO 2 C‐RP folgt eine Transaminierung, die mithilfe von Glutamat AFP liefert. RosB wurde auch von einer anderen Forschungsgruppe als Schlüsselenzym der Roseoflavinbiosynthese beschrieben . Allerdings wurde in dieser Arbeit die Notwendigkeit von Thiamin zur Bildung von HO 2 C‐RP bzw.…”

“…O 2 reagiert mit RP an der C8‐Methylgruppe, das daraus entstehende Hydroperoxyflavin wird protoniert und dissoziiert sofort zu OHC‐RP und OH − . Dieser Reaktionsweg würde übrigens auch eine Wasserstoffperoxidbildung umgehen, die weder von uns noch von anderen beobachtet wurde . Ein weiteres aktiviertes Wassermolekül liegt in unmittelbarer Nähe von C8 und könnte für die Bildung von HO 2 C‐RP aus OHC‐RP verantwortlich sein.…”