Ebony, a novel nonribosomal peptide synthetase for beta-alanine conjugation with biogenic amines in Drosophila.
Using Ebony protein either expressed in Escherichia coli or in Schneider S2 cells, we provide evidence for its substrate specificity and reaction mechanism. Ebony activates beta-alanine to aminoacyladenylate by an adenylation domain and covalently attaches it as a thioester to a thiolation domain in a nonribosomal peptide synthetase (NRPS) related mechanism. In a second reaction, biogenic amines act as external nucleophiles on beta-alanyl-S-pantetheine-Ebony, thereby releasing in a fast reaction the dipeptide (peptidoamine) in a process that is novel in higher eucaryotes. Therefore, we define Ebony as a beta-alanyl-biogenic amine synthetase. Insight into the reaction mechanism stems from mutational analysis of an invariant serine that disclosed Ebony as a multienzyme with functional analogy to the starting modules of NRPSs. In light of a putative biogenic amine-deactivating capacity, Ebony function in the nervous system must be reconsidered. We propose that in the Drosophila eye Ebony is involved in the transmission process by inactivation of histamine through beta-alanyl conjugation.
Richardt
Arnd
A
Fakultät für Chemie, AG Molekulare Zellbiochemie, Ruhr-Universität Bochum, Universitätsstrasse 150, 44780 Bochum, Germany.
Kemme
Tobias
T
Wagner
Stefanie
S
Schwarzer
Dirk
D
Marahiel
Mohamed A
MA
Hovemann
Bernhard T
BT
D000409:Alanine / Q000737:chemistry; D000818:Animals; D002460:Cell Line; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000378:metabolism; D004331:Drosophila melanogaster; D004926:Escherichia coli / Q000378:metabolism; D005091:Exons; D006632:Histamine / Q000737:chemistry; D007700:Kinetics; D008954:Models, Biological; D008956:Models, Chemical; D008957:Models, Genetic; D009420:Nervous System / Q000378:metabolism; D010453:Peptide Synthases / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011755:Pyrophosphatases / Q000378:metabolism; D015091:beta-Alanine / Q000737:chemistry*
D018076:DNA, Complementary; D010455:Peptides; D015091:beta-Alanine; D006632:Histamine; D011755:Pyrophosphatases; C019757:ATP pyrophosphohydrolase; D010453:Peptide Synthases; D000409:Alanine