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(Last
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The biosynthesis of the gerberin aglycone by 2-pyrone synthase (2PS)
Key reference: Eckermann et al. (1998) This was at the time of discovery quite a new finding for a physiological role in the family of CHS-related proteins: A type III PKS which used acetyl-CoA as starter, carried out two condensations, and released a pyrone. The reaction is shown below. One of the at first confusing properties of the enzyme is that it can decarboxylate malonyl-CoA to acetyl-CoA, thus synthesizing its own substrate from the standard chain extender. Why was that confusing initially? A typical negative control in these assays is that one omits the predicted starter substrate, in this case: acetyl-CoA. If you do that in such a case, you'll get despite of that rather high enzyme activities: the enzyme simple makes its own substrate. Studies with other type III PKS later showed that almost all of them can decarboxylate malonyl-CoA (see: Eckermann et al., 2003), but this usually is not noticed because in most cases acetyl-CoA or malonyl-CoA are not substrates for most of the plant type III PKS. Work in the last few years has shown, however, that there plant other enzymes using acetyl-CoA/malonyl-CoA as starter molecules. Interestingly, most if not all of them are enzymes carrying out four, five, six, or even seven
(a, b) condensation reactions! Acetyl-CoA or/and malonyl-CoA are also typical substrates for bacterial type III PKS (more...).
 Type III PKS in the biosynthesis of gerberin and parasorboside, via triacetic acid lactone (a pyrone).
The protein is a pyrone synthase that uses acetyl-CoA as starter substrate and performs two condensation reactions. The enzyme also decarboxylates the chain extender malonyl-CoA to acetyl-CoA, and thus can synthesize its own starter substrate. Also take a look at some interesting aspect of the fact that the 2PS is not only active with acetyl-CoA, but also with other small, hydrophobic CoA-esters, e.g. benzoyl-CoA, synthesizing the backbone of molecules that might be of medical interest:
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References
Eckermann, S., Schröder, G., Schmidt, J., Strack, D., Edrada, R.A., Helariutta, Y., Elomaa, P., Kotilainen, M., Kilpeläinen, I., Proksch, P., Teeri, T.H. and Schröder, J.: New pathway to polyketides in plants. Nature (London) 396, 387-390 (1998).
The repertoire of secondary metabolism (involving the production of compounds not essential for growth) in the plant kingdom is enormous, but the genetic and functional basis for this diversity is hard to analyse as many of the biosynthetic enzymes are unknown. We have now identified a key enzyme in the ornamental plant Gerbera hybrida (Asteraceae) that participates in the biosynthesis of compounds that contribute to insect and pathogen resistance. Plants transformed with an antisense construct of gchs2, a complementary DNA encoding a previously unknown function, completely lack the pyrone derivatives gerberin and parasorboside. The recombinant plant protein catalyses the principal reaction in the biosynthesis of these derivatives: GCHS2 is a polyketide synthase that uses acetyl-CoA and two condensation reactions with malonyl-CoA to form the pyrone backbone of the natural products. The enzyme also accepts benzoyl-CoA to synthesize the backbone of substances that have become of interest as inhibitors of the HIV-1 protease. GCHS2 is related to chalcone synthase (CHS) and its properties define a new class of function in the protein superfamily. It appears that CHS-related enzymes are involved in the biosynthesis of a much larger range of plant products than was previously realized.
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Eckermann, C., Schröder, G., Eckermann, S., Strack, D., Schmidt, J., Schneider, B., and Schröder, J.: Stilbenecarboxylate biosynthesis: a new function in the family of chalcone synthase-related proteins. Phytochemistry 62, 271-286 (2003).
Chalcone (CHS), stilbene (STS) synthases, and related proteins are key enzymes in the biosynthesis of many secondary plant products. Precursor feeding studies and mechanistic rationalization suggest that stilbenecarboxylates might also be synthesized by plant type III polyketide synthases; however, the enzyme activity leading to retention of the carboxyl moiety in a stilbene backbone has not yet been demonstrated. Hydrangea macrophylla L. (Garden Hortensia) contains stilbenecarboxylates (hydrangeic acid and lunularic acid) that are derived from 4-coumaroyl and dihydro-4-coumaroyl starter residues, respectively. We used homology-based techniques to clone CHS-related sequences, and the enzyme functions were investigated with recombinant proteins. Sequences for two proteins were obtained. One was identified as CHS. The other shared 65-70% identity with CHSs and other family members. The purified recombinant protein had stilbenecarboxylate synthase (STCS) activity with dihydro-4-coumaroyl-CoA, but not with 4-coumaroyl-CoA or other substrates. We propose that the enzyme is involved in the biosynthesis of lunularic acid. It is the first example of a STS-type reaction that does not lose the terminal carboxyl group during the ring folding to the end product. Comparisons with CHS, STS, and a pyrone synthase showed that it is the only enzyme exerting a tight control over decarboxylation reactions. The protein contains unusual residues in positions highly conserved in other CHS-related proteins, and mutagenesis studies suggest that they are important for the structure or/and the catalytic activity. The formation of the natural products in vivo requires a reducing step, and we discuss the possibility that the absence of a reductase in the in vitro reactions may be responsible for the failure to obtain stilbenecarboxylates from substrates like 4-coumaroyl-CoA.
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