Acyl carrier protein domain, polyketide

Modular polyketide synthases synthesize nascent polyketide scaffolds in an assembly line-like process, with each module participating in a single round of chain elongation and modification. At a minimum, all modules posses a B-keto synthase catalytic domain, an acyl transferase domain, and an acyl carrier protein domain. In general, specialized modules exist at the N-terminus of specific... 

K.J. Weissman, H. Hong, B. Popovic, F. Meersman, Evidence for a protein-protein interaction motif on an acyl carrier protein domain from a modular polyketide synthase. Chem. Biol. 13, 625-636 (2006)... 

V.Y. Alekseyev, C.W. Liu, D.E. Cane, J.D. Puglisi, C. Khosla, Solution structure and proposed domain-domain recognition interface of an acyl carrier protein domain from a modular polyketide synthase. Protein Sci. 16, 2093-2107 (2007)... 

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... 

Figure 21-11 Catalytic domains within three polypeptide chains of the modular polyketide synthase that forms 6-deoxyerythronolide B, the aglycone of the widely used antibiotic erythromycin. The domains are labeled as for fatty acid synthases AT, acyltransferase ACP, acyl carrier protein KS, 3-ketoacyl-ACP synthase KR, ketoreductase DH, dehydrase ER, enoylreductase TE, thioesterase. After Pieper et al.338 Courtesy of Chaitan Khosla.

Each synthetase module contains three active site domains The A domain catalyzes activation of the amino acid (or hydroxyacid) by formation of an aminoacyl- or hydroxyacyl-adenylate, just as occurs with aminoacyl-tRNA synthetases. However, in three-dimensional structure the A domains do not resemble either of the classes of aminoacyl-tRNA synthetases but are similar to luciferyl adenylate (Eq. 23-46) and acyl-CoA synthetases.11 The T-domain or peptidyl carrier protein domain resembles the acyl carrier domains of fatty acid and polyketide synthetases in containing bound phos-phopantetheine (Fig. 14-1). Its -SH group, like the CCA-terminal ribosyl -OH group of a tRNA, displaces AMP, transferring the activated amino acid or hydroxy acid to the thiol sulfur of phosphopan-tetheine. The C-domain catalyzes condensation (peptidyl transfer). The first or initiation module lacks a C-domain, and the final termination module contains an extra termination domain. The process parallels that outlined in Fig. 21-11.1... 

R Dieckmann, H von Dohren. Structural model of acyl carrier domains in integrated biosynthetic systems forming peptides, polyketides and fatty acids based on analogy to the E. coli acyl carrier protein. In RH Baltz, GD Hegeman, PL Skatrud, eds. Developments in Industrial Microbiology. Fairfax, VA Society for Industrial Microbiology, 1997, pp 79-87. 

Figure 5 Domain organization of the erythromycin polyketide synthase. Putative domains are represented as circles and the structural residues are ignored. Each module incorporates the essential KS, AT, and ACP domains, while all but one include optional reductive activities. AT, acyltransferase ACP, acyl carrier protein KS, (3-ketoacyl synthase KR, P-ketoacyl reductase DH, dehydratase ER, enoyl reductase TE, thioesterase.

J-A Chuck, M McPherson, H Huang, JR Jacobsen, C Khosla, DE Cane. Molecular recognition of diketide substrates by a P-keto-acyl carrier protein synthase domain within a bimodular polyketide synthase. Chem Biol 4 757-766, 1997. 

SF Haydock, JF Aparicio, I Molnar, T Schwecke, LE Khaw, A Konig, AFA Marsden, IS Galloway, J Staunton, PF Leadlay. Divergent sequence motifs correlated with the substrate specificity of (methyl)malonyl-CoA acyl carrier protein trans-acylase domains in modular polyketide synthases. FEBS Lett 374 246-248, 1995. 

Haydock SF, Aparicio JF, Mobiar I, Schwecke T, Khaw LE, Konig A, Marsden AF, Galloway IS, Staunton J, Leadlay PF (1995) Divergent Sequence Motifs Correlated with the Substrate Specificity of (Methyl) mMalonyl-CoA Acyl Carrier Protein Transacylase Domains in Modular Polyketide Synthases. FEBS Lett 374 246... 

According to the now widely accepted model of Katz and coworkers (Fig. 7), the loading acyltransferase (AT-L) domain at the NHj-terminal of DEBS 1 initiates the polyketide chain-building process by transferring the propionyl-CoA primer unit via the pantetheinyl residue of the first acyl carrier protein... 

Crawford JM, Dancy BCR, Hill EA, Udwary D, Townsend CA. Identification of a starer unit-acyl carrier protein transacylase domain in an iterative type 1 polyketide synthase. Proc. Natl. Acad. Sci. U.S.A. 2006 103 16728-16733. 

Figure 1 Hypothetical pentaketide biosynthetic system, which illustrates the enzymatic logic of type I modular polyketide synthases (PKSs) and the catalytic role of acyl transferase (AT) domains. Each AT domain selects substrates from the cellular pool and tethers them as thioesters to acyl carrier protein (ACP) domains. In a typical PKS module, the AT and ACP domains are present in all modules. The ketosynthase (KS) domain is present in all chain extension modules. The dehydratase (DH), enoyl reductase (ER), and ketoreductase (KR) domains are optional domains. The final thioesterase (TE) domain catalyzes the release of the product from the PKS.

Since the PKS (polyketide synthase) gene cluster for actinorhodin (act), an antibiotic produced by Streptomyces coelicolor[ 109], was cloned, more than 20 different gene clusters encoding polyketide biosynthetic enzymes have been isolated from various organisms, mostly actinomycetes, and characterized [98, 100]. Bacterial PKSs are classified into two broad types based on gene organization and biosynthetic mechanisms [98, 100, 102]. In modular PKSs (or type I), discrete multifunctional enzymes control the sequential addition of thioester units and their subsequent modification to produce macrocyclic compounds (or complex polyketides). Type I PKSs are exemplified by 6-deoxyerythronolide B synthase (DEBS), which catalyzes the formation of the macrolactone portion of erythromycin A, an antibiotic produced by Saccharopolyspora erythraea. There are 7 different active-site domains in DEBS, but a given module contains only 3 to 6 active sites. Three domains, acyl carrier protein (ACP), acyltransferase (AT), and P-ketoacyl-ACP synthase (KS), constitute a minimum module. Some modules contain additional domains for reduction of p-carbons, e.g., P-ketoacyl-ACP reductase (KR), dehydratase (DH), and enoyl reductase (ER). The thioesterase-cyclase (TE) protein is present only at the end of module 6. 

Modular PKSs are large multifunctional enzymes. Active sites (domains) within these enzymes ketosynthases (KS), acyltransferases (AT), dehydratases (DH), enoyl reductases (ER), ketoreductases (KR), acyl carrier proteins (AGP) and thioesterases (TE) are organized into modules such that each module catalyzes the stereospecific addition of a new monomer onto a growing polyketide chain and also sets the reduction level of the carbon atoms of the resulting intermediate [70]. In 1994, the heterologous expression of the complete erythromycin polyketide synthase was accomplished. The recombinant... 

Fig. 3. Phosphopantetheinylation of the acyl carrier protein (AGP) domain of a polyketide synthase. In order to be active, polyketide synthases must be post-translationally modified by a family of enzymes called phosphopantetheine transferases (PPTases). These enzymes transfer the 4 -phospho-pantetheine arm of Coenzyme A to an active site serine residue in the AGP...

---