Multifunctional enzyme protein

The polyketide synthases and peptide synthetases catalyze an unusually large number of reactions. 6-Methylsalicylic acid synthase, which produces 6-methylsalicylic acid from acetyl Co A, malonyl Co A and NADPH, for instance, carries out a total of thirteen reactions. The peptide antibiotic synthetases activate each of the amino acids involved in the formation of the peptides by a two step mechanism at a specific peripheral domain of the enzyme protein and catalyze also linkage of the activated amino acids by sequential transpeptidation. In the case of gramicidin S synthetase for example, there are twenty-one different reactions carried out by two multifunctional enzyme proteins. 

Compound 508 (PNRI-299), a small molecule /3-strand mimetic template compound, is active as an inhibitor of the multifunctional activating protein factor 1 (AP-1). PNRI-299 has demonstrable effects on the reduction of API-driven transcription and beneficial pharmacological effects in a mouse asthma model. The compound affects the expression of leukotriene C4 (LTC4) synthase, a crucial enzyme for the formation of the cysteinyl leukotrienes <2003PNA1169, 2004MI211>. 

A closely related E. coli protein is a 79-kDa multifunctional enzyme that catalyzes four different reactions of fatty acid oxidation (Chapter 17). The amino-terminal region contains the enoyl hydratase activity.32 A quite different enzyme catalyzes dehydration of thioesters of (3-hydroxyacids such as 3-hydroxydecanoyl-acyl carrier protein (see Eq. 21-2) to both form and isomerize enoyl-ACP derivatives during synthesis of unsaturated fatty acids by E. coli. Again, a glutamate side chain is the catalytic base but an imidazole group of histidine has also been implicated.33 This enzyme is inhibited irreversibly by the N-acetylcysteamine thioester of 3-decynoic acids (Eq. 13-8). This was one of the first enzyme-activated inhibitors to be studied.34... 

Polyketides are made by the sequential activity of domains of large, multifunctional enzymes called polyketide synthases (PKSs) (Fig. 6a and b). Polyketides are formed by the condensation and modification of acyl units derived from acyl-CoA precursors. Domains are organized in modules and each module carries out the series of steps necessary for one cycle of polyketide chain elongation. A single protein can have more than one module, and several different proteins together can make up a PKS. The number of modules determines the size of the polyketide. A growing polyketide chain is tethered to the enzyme as a thiol ester and moves sequentially from the N- to the C-terminus of a module, lengthened by two carbon units per module. The first module in a PKS... 

The animal fatty acid synthase (FAS EC 2.3.1.85) is one of the most complex multifunctional enzymes that have been characterized, as this single polypeptide contains all the catalytic components required for a series of 37 sequential transactions (Smith, 1994). The animal FAS consists of two identical polypeptides of approximately 2500 amino acid residues (MW, ca. 270 kDa), each containing seven catalytic subunits (1) ketoacylsynthase, (2) malonyl/acetyl transferase, (3) dehydrase, (4) enoyl reductase, (5) (3-kcto reductase, (6) acyl carrier protein (ACP), and (7) thioesterase. Although some components of the complex are able to carry out their respective catalytic steps in the monomeric form, only in the FAS dimer do the subunits attain conformations that facilitate coupling of the individual reactions of fatty acid synthesis to occur (Smith et al., 2003). 

One of the most exciting areas in the field of multifunctional enzyme systems is the synthesis of a wide array of organic molecules by polyketide and nonribosomal protein synthetases. These enzymes are generally characterized by multiple subunits which themselves consist of individual domains with distinct enzymatic activities (Fig. 9.11a). The range of natural products synthesized by these mega-synthetases includes a considerable number of important antibiotics, antifungals, antitumor and cholesterol-lowering compounds, immunosuppressants, and siderophores. 

Dissection of Multifunctional Molecules. Protein molecules generally have more than one antigenic site and occasionally more than one distinctly diflFerent biologically active site. Whenever these sites are discretely separated spatially and are joined by only one or a few interconnecting peptide chains, proteolytic enzymes may separate the sites whil preserving biological activities. Proteolytic scission is most successful when the molecule is elongated or composed of more than one polypeptide chain. 

Agouron began with consideration of the structure of the complex between the E. coll enzyme and 5-deazatetrahydrofolate (77). An active and soluble fragment of a multifunctional human protein that contained the GARFT activity was provided by recombinant approaches (78), and its structure was also solved (79) in complex with novel inhibitors. Comparison of the two structures subsequently validated the use of the bacterial enzyme as a model for the human GARFT. The design of novel inhibitors also relied on previous studies of the structure-activity relationships (SAR) for substitutents around the core... 

The inhibition of multifunctional enzymes can also have therapeutic interest. The 26S proteasome is a mul-ticatalytic intracellular protease complex expressed in eukaryotic cells. This complex is responsible for selective degradation of intracellular proteins that are responsible for cell proliferation, growth, regulation of apoptosis and transcription of genes. Thus, proteasome inhibition is a potential treatment option for cancer and diseases due to aberrant inflammation conditions. Bortezomib and PS-519 are the first proteasome inhibitors that have entered clinical trials. In multiple myeloma, both the FDA (United States... 

Many eukaryotic multienzyme complexes are multifunctional proteins in which different enzymes are linked covalently. An advantage of this arrangement is that the synthetic activity of different enzymes is coordi nated. In addition, intermediates can be efficiently handed from one active site to another without leaving the assembly. Furthermore, a complex of covalently joined enzymes is more stable than one formed by noncovalent attractions. Each of the component enzymes is recognizably homologous to its bacterial counterpart. It seems likely that multifunctional enzymes sudi as fatty acid synthase arose in eukaryotic evolution by fusion of the individual genes of evolutionary ancestors,... 

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