Nonribosomal peptide synthase NRPS

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al. 1997 9 Mayes et al. 1993 10 Shick et al. 1999 11 Richards et al. 2006 12 Bouarab et al. 2004 13 Pelletreau et al., unpublished data 14 Dittman and Weigand 2006 15 Rein and Barrone 1999 Empty columns imply no direct evidence of these enzymes from these systems... 

Fig. 9.11. Organizational overview and summary of protein engineering efforts on the poly-ketide synthetase (PKS) and nonribosomal peptide synthetase (NRPS) framework, a) In both systems, the complete synthetases consist of multiple subunits encoded on individual genes. The subunits themselves are divided in modules which each catalyze the addition of one acylbuilding block (PKS) or amino acid (NRPS). The minimal module for PKSs is made up of three domains - a keto synthase (KS), an acyl transferase (AT), and an acyl carrier protein (ACP). In addition, modules can contain up to three modifying domains to derivatize (S-carbons on the...

Several of these genes vere deduced to program for polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) enzymes as well as novel tailoring enzymes, including those that create the various unusual functional groups in jamaicamide A. 

Many important therapeutics, in use in clinics today, are biosynthesized by the nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) paradigm. For example, many of the antibiotics (penicillin, cephalosporin, vancomycin, erythromycin, etc.), immunosuppressors (cyclosporine, rapamycin), antiviral agents (luzopeptin A), antitumor agents (bleomycin), and toxins (thaxtomin) are NRPS and PKS derived.20-22 Figure 1 displays a small selection of natural products that are NRPS and PKS derived and illustrates the diversity of molecular structures generated by these biosynthetic paradigms. 

Figure 6 Organization of the polyketide synthase (PKS)/nonribosomal peptide synthetase (NRPS) system for which ketosynthase (KS) and colinearity rule-based predictions have assisted the discovery of the thailandamides from Burkholderia thailandensis. The different proteins are highlighted in different colors and each module is boxed. The experimentally determined structure of thailandamide A is shown at the bottom of the figure.

The biosynthesis of pyridomycin was first studied by Ogawara. Isotope labeling studies demonstrated that it might involve the assembly of the backbone by a hybrid nonribosomal peptide synthetase (NRPS)/polyketide synthase (PKS) system using 3-hydroxypicolinic acid (3-HPA) as the starting unit (Figure 6.15) [67]. 

Kalkitoxin must be derived from a mixed polyketide/nonribosomal peptide synthase pathway, hi spite of intensive research on cyanobacterial PKS/NRPS, the kalkitoxin biosynthesis gene was not disclosed. 

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. 

Phosphopantetheine tethering is a posttranslational modification that takes place on the active site serine of carrier proteins - acyl carrier proteins (ACPs) and peptidyl carrier proteins (PCPs), also termed thiolation (T) domains - during the biosynthesis of fatty acids (FAs) (use ACPs) (Scheme 23), polyketides (PKs) (use ACPs) (Scheme 24), and nonribosomal peptides (NRPs) (use T domain) (Scheme 25). It is only after the covalent attachment of the 20-A Ppant arm, required for facile transfer of the various building block constituents of the molecules to be formed, that the carrier proteins can interact with the other components of the different multi-modular assembly lines (fatty acid synthases (FASs), polyketide synthases (PKSs), and nonribosomal peptide synthetases (NRPSs)) on which the compounds of interest are assembled. The structural organizations of FASs, PKSs, and NRPSs are analogous and can be divided into three broad classes the types I, II, and III systems. Even though the role of the carrier proteins is the same in all systems, their mode of action differs from one system to another. In the type I systems the carrier proteins usually only interact in cis with domains to which they are physically attached, with the exception of the PPTases and external type II thioesterase (TEII) domains that act in trans. In the type II systems the carrier proteins selectively interact... 

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