BIOSYNTHESIS OF HYPERICINS AND HYPERFORINS IN HYPERICUM PERFORATUM L. (ST. JOHN’S WORT) – PRECURSORS AND GENES INVOLVED, ACTA UNIVERSITATIS OULUENSIS A Scientiae Rerum Naturalium 564

Hypericum perforatum L. (St. John’s wort) is a medicinal plant widely utilized for the treatmentof depression. The antidepressant activity is mainly attributed to the phenolic compoundshypericins and hyperforins, which also have a wide range of other pharmacologically interestingproperties. The biosynthetic routes leading to hypericins and hyperforins are poorly understood,although a polyketide pathway including type III polyketide synthases (PKSs) has been suggestedto be involved. Furthermore, a gene called hyp-1 is assumed to attend to the final stages of thehypericin biosynthesis. In the present work, the biosynthesis of hypericins and hyperforins in H.perforatum was further studied by focusing on the elucidation of the precursors and genesinvolved. \nThe incorporation of isotopically labelled branched-chain amino acids into hyperforins wasinvestigated as well as the possibilities to enhance the production of hyperforins in H. perforatumin vitro cultures by feeding them with amino acid precursors. Furthermore, two novel cDNAsencoding for type III PKSs were isolated from H. perforatum. The functions of these new genes,designated HpPKS1 and HpPKS2, as well as the role of hyp-1 were elucidated by comparing theirexpression with the levels of hypericins and hyperforins in H. perforatum tissues. The enzymaticactivity of the recombinant HpPKS2 protein was also analyzed. To study Hyp-1 at a protein level,a protein extraction method was optimized for tissues of Hypericum species. \nThe results show the incorporation of valine and isoleucine into the acyl side chain ofhyperforin and adhyperforin, respectively. Through the biotransformation of the amino acidprecursors, it is possible to enhance the levels of adhyperforin, but not hyperforin, in H.perforatum shoot cultures, which demonstrates the tight regulation of the hyperforin biosynthesis.A correlation between HpPKS1 expression and hyperforins was detected in H. perforatum tissues.The localization of HpPKS2 mRNA in dark glands in which hypericins accumulate as well as theoctaketide synthase activity of the recombinant HpPKS2 suggest that HpPKS2 is associated withpossible co-operating tailoring enzymes in the biosynthesis of hypericins. The presence of bothhyp-1 mRNA and Hyp-1 protein in distinct places compared with hypericins in H. perforatumtissues does not support the idea that Hyp-1 would be involved in the biosynthesis of hypericinsin dark glands, although mobility of the Hyp-1 protein was shown to be possible. \nThe present thesis extends knowledge about the biosynthesis of hypericins and hyperforins inH. perforatum by providing new candidate genes for their biosynthesis and by identifyingprecursors for hyperforins. Moreover, new information was obtained about the role of hyp-1 in H.perforatum.

ISBN-10:
978-951-42-6309-5
Kieli:
eng.
Sivumäärä:
140 s.
Tekijät:
KARPPINEN KATJA
Tuotekoodi 014007
21,00 €