University of Kentucky
Enzymatic Evidence for a Revised Congocidine Biosynthetic Pathway
Institution
University of Kentucky
Faculty Advisor/ Mentor
Sylvie Garneau-Tsodikova
Abstract
Nonribosomal peptides are proteins that are assembled through a chain of other proteins called nonribosomal peptide synthetases, many of which are composed of adenylation (A), thiolation (T), and condensation (C) units corresponding to function. Naturally produced pyrrolamides, such as congocidine, are nonribosomal peptides that bind to the minor groove of DNA in a sequence-specific manner, which confers anticancer activity. Considerable efforts have been made to increase this ability and overcome toxicity of pyrrolamide analogues. The majority of attempts to delineate the biosynthetic machinery, the order and method of protein function, during pyrrolamide assembly have been genetic methods. We discussed biochemical characterization of four essential proteins in congocidine formation: the adenylation-thiolation (A-T) di-domain Cgc18(1-610) and its MbtH-like partner SAMR0548, the AMP-binding enzyme Cgc3*, and the T domain Cgc19. A substrate is a molecule that an enzyme specifically binds and alters to catalyze substrate formation into a product. AMP stands for adenosine monophosphate, a molecule involved in energy transfer. We reported revised substrate specificities of Cgc18(1- 610) and Cgc3*, biochemical evidence of the AMP-binding activity of Cgc3*, and loading of 4- acetamidopyrrole-2-carboxylic acid onto Cgc19. Based on these biochemical studies, we suggest a revised congocidine biosynthetic pathway.
Enzymatic Evidence for a Revised Congocidine Biosynthetic Pathway
Nonribosomal peptides are proteins that are assembled through a chain of other proteins called nonribosomal peptide synthetases, many of which are composed of adenylation (A), thiolation (T), and condensation (C) units corresponding to function. Naturally produced pyrrolamides, such as congocidine, are nonribosomal peptides that bind to the minor groove of DNA in a sequence-specific manner, which confers anticancer activity. Considerable efforts have been made to increase this ability and overcome toxicity of pyrrolamide analogues. The majority of attempts to delineate the biosynthetic machinery, the order and method of protein function, during pyrrolamide assembly have been genetic methods. We discussed biochemical characterization of four essential proteins in congocidine formation: the adenylation-thiolation (A-T) di-domain Cgc18(1-610) and its MbtH-like partner SAMR0548, the AMP-binding enzyme Cgc3*, and the T domain Cgc19. A substrate is a molecule that an enzyme specifically binds and alters to catalyze substrate formation into a product. AMP stands for adenosine monophosphate, a molecule involved in energy transfer. We reported revised substrate specificities of Cgc18(1- 610) and Cgc3*, biochemical evidence of the AMP-binding activity of Cgc3*, and loading of 4- acetamidopyrrole-2-carboxylic acid onto Cgc19. Based on these biochemical studies, we suggest a revised congocidine biosynthetic pathway.