Enzymes biosynthesis associated

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. 

Rubber is synthesized by plants via a side branch of the isoprenoid pathway by the enzyme rubber transferase (dy-prenyl transferase systematic name poly-dy-polyprenyl-diphosphate isopentenyl-diphosphate polyprenylcistransferase EC 2.5.1.20). Surprisingly, although this process has been studied for decades, due to the labile nature of the rubber transferase and the fact that it is a membrane-associated enzyme present in relatively low abundance, the identification of its protein subunits remain elusive. For some recent reviews on rubber biosynthesis, please refer to [248-251]. 

Figure 2.12 A hypothetical view of compartmentation of indole alkaloid biosynthesis in Catharanthus roseus. Enzymes located with dashed arrows are hypothetical and circles indicate membrane associated enzymes (after Meijer et at, 1 993b). Cl OH, geraniol-1 0-hydroxylase NMT, 5-adenosyl-L-methionine 11 -methoxy 2,16-dihydro-16-hydroxytabersonine N-methyltransferase DAT, acetylcoenzyme A deacetylvindoline 1 7-0-acetyltransferase OHT, 2-oxyglutarate-dependent dioxygenase SSpC, strictosidine-((3)-glucosidase SSS, strictosidine synthase.

ACP is a central component of type II PKS and is involved in possibly all reactions of bacterial aromatic polyketide biosynthesis. The starter unit (before its transfer to the KS subunit), the extender units, the growing poly-)5-ketone intermediates, as well as the full length linear poly-)8-ketone product, are covalently bound to ACP in a thioester linkage to the terminal sulfhydryl of the 4 -phos-phopantetheine prosthetic group (see Sect. 3.1.1). These ACP thioesters must have been recognized as substrates by type II PKSs and other associated enzymes such as KR, CYC, or ARC to build and process the poly- -ketone intermediates into aromatic polyketides. 

Fatty acid synthase. Fatty acid biosynthesis in plants occurs primarily in the plastids and is dependent upon the existence of the fatty acid synthase (or synthetase) II complex (FAS) (Harwood, 1988 Stumpf, 1989). This complex, unlike the system present in animals, is discontinuous (Shimakata and Stumpf, 1982a). Thus it has been possible to isolate and study the associated enzyme activities independently. Purification of each activity to homogeneity can then allow amino acid sequence analysis, cloning and elucidation of the gene, and subsequent application in the genetic engineering of plants. 

A number of enzymatic processes associated with hydroxy-lations, such as NADH2 / NAD redox interactions, showed significant catalysis when AA was added These activities were attributed to primarily oxidation - reduction involving the components of the ascorbic system. But recent work has revealed that in a number of reactions the effect may be Interpreted not in terms of direct redox activities, but rather in terms of the effect of AA on enzymatic aggregation and / disaggregation as well as in contribution to biosynthesis of enzyme components ... 

Tyrosine hydroxylase and dopamine- -hydroxylase occur only in those cells which synthesize catecholamines (adrenal medullary cells and adrenergic neurons). When the sympathetic innervation of peripheral organs is destroyed (surgically, immunologically or chemically) these enzyme activities disappear. This is not true of DOPA decarboxylase, which occurs in many cells apart from those involved in catecholamine biosynthesis. This enzyme, for example, is also found in S-hydroxy-tryptamine-containing cells, and in the kidney, liver and certain glial cells associated with cerebral blood vessels. 

Cytotoxicity occurs as a result of the molecular interactions of an alkaloid with one or several important targets present in a cell (Figures 1 and 2). The main targets include DNA, RNA, and the associated enzymes and processes (i.e., replication, repair, transcription, DNA polymerase, RNA polymerase, reverse transcriptase, repair enzymes, topoisomerase, telomerase), protein biosynthesis, protein conformation, biomembranes, and membrane proteins (for reviews, see (10,11,16)). 

Biosynthesis of Tea Flavonoids. The pathways for the de novo biosynthesis of flavonoids in both soft and woody plants (Pigs. 3 and 4) have been generally elucidated and reviewed in detail (32,51). The regulation and control of these pathways in tea and the nature of the enzymes involved in synthesis in tea have not been studied exhaustively. The key enzymes thought to be involved in the biosynthesis of tea flavonoids are 5-dehydroshikimate reductase (52), phenylalanine ammonia lyase (53), and those associated with the shikimate/arogenate pathway (52). At least 13 enzymes catalyze the formation of plant flavonoids (Table 4). 

All prostaglandins are cyclopentanoic acids derived from arachidonic acid. The biosynthesis of prostaglandins is initiated by an enzyme associated with the endoplasmic reticulum, called prostaglandin endoperoxide synthase, also known as cyclooxygenase. The enzyme catalyzes simultaneous oxidation and cyclization of arachidonic acid. The enzyme is viewed as having two distinct activities, cyclooxygenase and peroxidase, as shown in Figure 25.28. 

For the sake of study, the biosynthesis of carotenoid plant pigments can be divided into parts involving enzymes and their associated activities as listed in Table 5.3.1 and further detailed in Figure 5.3.1 through Figure 5.3.4. Some of the parts have common enzymatic mechanisms and may also be in distinct subcellular compartments such as cytoplasm, endoplasmic reticulum, or plastid thylakoid. 

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