The invention relates generally to materials and methods for biosynthesising quillaic acid in a host by expressing heterologous nucleotide sequences in the host each of which encodes a polypeptide which in combination have said QA biosynthesis activity. Example polypeptides include (i) a Beta-amyrin synthase; (ii) an enzyme capable of oxidising Beta-amyrin or an oxidised derivative thereof at the C-28 position to a carboxylic acid; (iii) an enzyme capable of oxidising Beta-amyrin or an oxidised derivative thereof at the C-16a position to an alcohol; and (iv) an enzyme capable of oxidising Beta-amyrin or an oxidised derivative thereof at the C-23 position to an aldehyde. Preferred nucleotide sequences are obtained from, or derived from, Q. saponaria.

An engineered microbial cell expressing a β-amyrin synthase, a cytochrome P450 reductase, a cytochrome P450 C28 oxidase, a cytochrome P450 C16 oxidase and a cytochrome C23 oxidase is used to make quillaic acid from β-amyrin.

The present invention relates inter alia to methods of biosynthetic production of QS-21, precursors and variants thereof, and to related aspects.

The invention relates generally to materials and methods for biosynthesising quillaic acid in a host by expressing heterologous nucleotide sequences in the host each of which encodes a polypeptide which in combination have said QA biosynthesis activity. Example polypeptides include (i) a Beta-amyrin synthase; (ii) an enzyme capable of oxidising Beta-amyrin or an oxidised derivative thereof at the C-28 position to a carboxylic acid; (iii) an enzyme capable of oxidising Beta-amyrin or an oxidised derivative thereof at the C-16 position to an alcohol; and (iv) an enzyme capable of oxidising Beta-amyrin or an oxidised derivative thereof at the C-23 position to an aldehyde. Preferred nucleotide sequences are obtained from, or derived from, Q. saponaria.

This invention relates to methods of producing triterpenoids using one or more of (i) Saponaria officinalis -amyrin synthase (SobAS) (ii) S. officinalis C28 oxidase (SoC28) (iii) S. officinalis C28C16 oxidase (SoC28C16)(iv) S. officinalis C23 oxidase (SoC23); (v) S. officinalis QA 3-O glucuronosyl transferase SoCSL; (vi) S. officinalis QA-GlcA SoC3Gal; (vii) S. officinalis QA-GlcA-Gal x SoC3Xy; (vii) S. officinalis QA-Tri fucosyl transferase SoC28Fu (ix) S. officinalis QA-TriF rhamnosyl transferase SoC28Rha (x) S. officinalis QA-TriFR xyl SoC28Xul1; (xi) S. officinalis QA-TriFRX xyl SoC28Xyl2; (xii) S. officinalis QA-TriFRXX quinovosyl SoGH1 and (xiii) A. officinalis QA-TroF(Q)RXX acetyl SoBAHD1 polypeptide. Methods, host cells, isolated polypeptides, nucleic acids, and plants are provided.