Nonribosomal peptide

It is likely that the biosynthesis of 113 is directed by a hybrid polyketide syn-thase/nonribosomal peptide synthetase enzyme system, as indicated in Figure 11.19. 

It is likely that the madurastatins are biosynthesized on a nonribosomal peptide synthetase, from salicylic acid as the starter acid. L-Serine is probably the precursor to the aziridine moiety, with epimerization occurring on the enzyme-bound amino acid as found for other nonribosomal peptides, with aziridine formation occurring at a late stage. Compounds 120 and 123 could therefore be biosynthetic precursors to 119 and 122, respectively. 

Figure 3.4 Improvement of the activity of chimeric NRPSs using directed evolution. (1) A heterologous A domain is swapped into an NRPS, typically resulting in a significant loss of synthetase activity. (2) A library of chimeric synthetase mutants is constructed in which the heterologous A domain has been diversified (for example, by error-prone PCR). (3) The library is subjected to an in vivo screen for production of the unnatural nonribosomal peptide derivative. (4) Clones showing improved production are characterized and subjected to further rounds of diversification and screening...

Fischbach, M.A. and Walsh, C.T. (2006) Assembly-line enzymology for polyketide and nonribosomal peptide antibiotics logic, machinery, and mechanisms. Chemical Reviews, 106, 3468—3496. 

Mootz, H.D., Schwarzer, D. and Marahiel, M.A. (2002) Ways of assembling complex natural products on modular nonribosomal peptide synthetases. Chembiochem A European Journal of Chemical Biology, 3, 490. 

Staunton, J. and Wilkinson, B. (2001) Combinatorial biosynthesis of polyketides and nonribosomal peptides. Current Opinion in Chemical Biology, 5, 159. 

Walsh, C.T. (2004) Polyketide and nonribosomal peptide antibiotics modularity and versatility. Science, 303, 1805. 

Sieber, S.A. and Marahiel, M.A. (2005) Molecular mechanisms underlying nonribosomal peptide synthesis approaches to new antibiotics. Chemical Reviews, 105, 715. 

Kopp, F. and Marahiel, M.A. (2007) Macrocyclization strategies in polyketide and nonribosomal peptide biosynthesis. Natural Product Reports, 24, 735. 

In this chapter, we will introduce an exciting class of natural product biosynthetic enzymes, the modular synthases, as well as their associated enzyme partners. We will discuss the use of metabolic engineering as a tool for small-molecule discovery and development, both through directed fermentation and combinatorial biosynthesis. In addition, we will review six classes of partner enzymes involved in the modification of polyketide (PK) and nonribosomal peptide (NRP) natural products. We believe that these enzymatic transformations hold great opportunities for synthetic chemists and will serve as the foundation for a new trend in both discovery and process chemistry. 

Cane, D.E. and Walsh, C.T. (1999) The parallel and convergent universes of polyketide synthases and nonribosomal peptide synthetases. Chemistry Biology, 6 (12), R319—R325. 

Trauger, J.W., Kohli, R.M. and Walsh, C.T. (2001) Cyclization of backbone-substituted peptides catalyzed by the thioesterase domain from the tyrocidine nonribosomal peptide synthetase. Biochemistry, 40 (24), 7092-7098. 

Duerfahrt, T., Eppelmann, K., Muller, R. andMarahiel, M.A. (2004) Rational design ofabimodular model system for the investigation of heterocyclization in nonribosomal peptide biosynthesis. Chemistry Biology, 11, 261-271. 

The biosynthetic hypothesis commences with the action of a nonribosomal peptide synthetase on natural amino acid precursors. While the incorporation of... 

Bacterial siderophores are typically small peptidic molecules, which contain side chains and functional groups that can provide high-affinity ligands for coordination of ferric ions. The structures of some siderophores are shown in Drechsel and Jung [142]. Siderophore synthesis occurs via enzymatic assembly by nonribosomal peptide synthetases [144], In bacteria, siderophore synthesis is... 

ENZ enzyme assays, SC structural composition, MM molecular methods, IL isotopic labeling, IF isotopic fractionation, INH inhibition studies, UNK unknown, LOX lipoxogenase, EPSP synthase 5-enolpyruvylshikimate-3-phosphate, SDH shikimate dehydrogenase, PAL phenylalanine ammonium lyase, PKS polyketide synthase, NRPS nonribosomal peptide synthase 1 Gerwick 1999 2 Liu et al. 1994 3 Boonprab et al. 2003 4 Cvejic and Rohmer 1999 5 Disch et al. 1998 6 Chikaraishi et al. 2006 7 Schwender et al. 2001 8 Schwender et al. 1997 9 Mayes et al. 1993 10 Shick et al. 1999 11 Richards et al. 2006 12 Bouarab et al. 2004 13 Pelletreau et al., unpublished data 14 Dittman and Weigand 2006 15 Rein and Barrone 1999 Empty columns imply no direct evidence of these enzymes from these systems... 

Earlier in this chapter, it was mentioned that many of the nonprotein amino acids are components of nonribosomal peptides. During such a biosynthesis, the peptide is attached to a carrier protein through a thioester bond, until chain termination occurs and the final product is released. The carrier protein is posttranslationally modified by the attachment of a phosphopantetheinyl group from coenzyme A. This step gives rise to the active carrier protein with a phosphopantetheine arm upon which amino acids are added to during NRPS. As an example, loading of isoleucine onto the carrier protein is depicted below (Scheme 5). Further details about nonribosomal peptide syntheses and enzymatic reactions can be found in Chapter 5.19. 

During the biosynthesis of nonribosomal peptides, there are two ways to incorporate the nonprotein amino acids. They can be incorporated either as a single unit or as an L-a-amino acid, which then undergoes structural modifications, while attached to the carrier protein. In the case of coronamic acid, L-rr//o-isoleucine is loaded onto the carrier protein and a unique biosynthetic pathway produces a cyclopropyl group containing a nonprotein amino acid. Specific examples of the biosynthesis of nonprotein amino acids will be discussed in the following sections. 

Many of the listed nonprotein amino acids have been identified as components of nonribosomal peptides from fungi and sponges. Hence, they tend to exhibit antifungal and antimicrobial properties, and in some cases potent cytotoxicity is observed. Specific details on known nonprotein amino acids are given in Table 1. 

Nonribosomal peptides from MIrazIrldIne - Cathepsin B Inhibitor 14, 30... 

A number of nonprotein amino acids with unsaturated side chains have been isolated. Many of these contain alkene side chains, but some alkyne side chains containing amino acids have also been identified. Nonprotein dehydroamino acids do not have an a-stereocenter these amino acids are still classified under this category. Dehydroamino acids are generally biosynthesized by the enzymatic elimination of a leaving group at the /3-carbon. For example, serine and threonine are enzymatically dehydrated to give dehydroalanine and dehydrobutyrine, respectively. A similar biosynthetic pathway is hypothesized for dehydroamino acids found in nonribosomal peptides, such as nodularins and microcystins. ... 

The glycopeptide antibiotics such as vancomycin and chloroeremomycin are complex nonribosomal peptides. One of the nonprotein amino acids found in these antibiotics is 4-hydroxyphenylglycine. The biosynthetic pathway of this nonprotein amino acid has been studied and prephenate was... 

Generally, most of the nonprotein amino acids containing aryl or functionalized aryl side chains are part of nonribosomal peptides isolated from hacteria, fungi, and sponges. These peptides exhibit interesting biological activities that include antimicrobial, antitumor, antifungal, and other inhibitory activities. Table 3 describes many of these types of amino acids. 

The nonprotein amino acid /3-A -methylamino-L-alanine (BMAA) is a neurotoxin found in various species of marine cyanobacteria. This nonprotein amino acid occurs both as a free amino acid and bound to proteins.Siderophores are secondary metabolites generally produced by bacteria under iron-deficient conditions. These molecules sequester and transport ferric ion via active transport in bacteria. Some known siderophores are nonribosomal peptides that contain nonprotein amino acids with terminal amines or hydroxylamine side chains, such as exochelin Many of the known nonprotein amino... 

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