Limits...
A Novel Class of Plant Type III Polyketide Synthase Involved in Orsellinic Acid Biosynthesis from Rhododendron dauricum

View Article: PubMed Central - PubMed

ABSTRACT

Rhododendron dauricum L. produces daurichromenic acid, the anti-HIV meroterpenoid consisting of sesquiterpene and orsellinic acid (OSA) moieties. To characterize the enzyme responsible for OSA biosynthesis, a cDNA encoding a novel polyketide synthase (PKS), orcinol synthase (ORS), was cloned from young leaves of R. dauricum. The primary structure of ORS shared relatively low identities to those of PKSs from other plants, and the active site of ORS had a unique amino acid composition. The bacterially expressed, recombinant ORS accepted acetyl-CoA as the preferable starter substrate, and produced orcinol as the major reaction product, along with four minor products including OSA. The ORS identified in this study is the first plant PKS that generates acetate-derived aromatic tetraketides, such as orcinol and OSA. Interestingly, OSA production was clearly enhanced in the presence of Cannabis sativa olivetolic acid cyclase, suggesting that the ORS is involved in OSA biosynthesis together with an unidentified cyclase in R. dauricum.

No MeSH data available.


The overall structures of M. sativa CHS2 and R. dauricum ORS. (A) The crystal structure of M. sativa CHS2 in complex with CoA (PDB: 1BQ6). The CoA molecule is depicted as blue sticks. (B) The model structure of ORS. The active site Cys164 in both enzymes is represented by a CPK model. The arrows indicate active site entrances. The inserted peptide sequences a–d in ORS are colored green and indicated by arrows.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5037138&req=5

Figure 4: The overall structures of M. sativa CHS2 and R. dauricum ORS. (A) The crystal structure of M. sativa CHS2 in complex with CoA (PDB: 1BQ6). The CoA molecule is depicted as blue sticks. (B) The model structure of ORS. The active site Cys164 in both enzymes is represented by a CPK model. The arrows indicate active site entrances. The inserted peptide sequences a–d in ORS are colored green and indicated by arrows.

Mentions: The primary structure comparison of ORS and CHS revealed several active site residue substitutions and unique peptide insertions. To assess the effects of these sequence changes on the protein structure, a homology model of ORS was constructed with the crystal structure of M. sativa CHS2 as a template, wherein the Ramachandran plot calculated that ∼96.7% of the amino acid residues were grouped in the energetically allowed regions. Figure 4B shows the overall structure of the ORS model, along with the crystal structure of M. sativa CHS2 (Figure 4A). Despite the relatively low sequence homology between the two enzymes, ORS adopts almost the same three-dimensional overall fold as that of M. sativa CHS2. In addition, all of the inserted peptide sequences in ORS were located on the protein surface, far from the active site cavity (Figure 4B), suggesting that these insertions minimally affect the protein folding and the catalytic activity.


A Novel Class of Plant Type III Polyketide Synthase Involved in Orsellinic Acid Biosynthesis from Rhododendron dauricum
The overall structures of M. sativa CHS2 and R. dauricum ORS. (A) The crystal structure of M. sativa CHS2 in complex with CoA (PDB: 1BQ6). The CoA molecule is depicted as blue sticks. (B) The model structure of ORS. The active site Cys164 in both enzymes is represented by a CPK model. The arrows indicate active site entrances. The inserted peptide sequences a–d in ORS are colored green and indicated by arrows.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5037138&req=5

Figure 4: The overall structures of M. sativa CHS2 and R. dauricum ORS. (A) The crystal structure of M. sativa CHS2 in complex with CoA (PDB: 1BQ6). The CoA molecule is depicted as blue sticks. (B) The model structure of ORS. The active site Cys164 in both enzymes is represented by a CPK model. The arrows indicate active site entrances. The inserted peptide sequences a–d in ORS are colored green and indicated by arrows.
Mentions: The primary structure comparison of ORS and CHS revealed several active site residue substitutions and unique peptide insertions. To assess the effects of these sequence changes on the protein structure, a homology model of ORS was constructed with the crystal structure of M. sativa CHS2 as a template, wherein the Ramachandran plot calculated that ∼96.7% of the amino acid residues were grouped in the energetically allowed regions. Figure 4B shows the overall structure of the ORS model, along with the crystal structure of M. sativa CHS2 (Figure 4A). Despite the relatively low sequence homology between the two enzymes, ORS adopts almost the same three-dimensional overall fold as that of M. sativa CHS2. In addition, all of the inserted peptide sequences in ORS were located on the protein surface, far from the active site cavity (Figure 4B), suggesting that these insertions minimally affect the protein folding and the catalytic activity.

View Article: PubMed Central - PubMed

ABSTRACT

Rhododendron dauricum L. produces daurichromenic acid, the anti-HIV meroterpenoid consisting of sesquiterpene and orsellinic acid (OSA) moieties. To characterize the enzyme responsible for OSA biosynthesis, a cDNA encoding a novel polyketide synthase (PKS), orcinol synthase (ORS), was cloned from young leaves of R. dauricum. The primary structure of ORS shared relatively low identities to those of PKSs from other plants, and the active site of ORS had a unique amino acid composition. The bacterially expressed, recombinant ORS accepted acetyl-CoA as the preferable starter substrate, and produced orcinol as the major reaction product, along with four minor products including OSA. The ORS identified in this study is the first plant PKS that generates acetate-derived aromatic tetraketides, such as orcinol and OSA. Interestingly, OSA production was clearly enhanced in the presence of Cannabis sativa olivetolic acid cyclase, suggesting that the ORS is involved in OSA biosynthesis together with an unidentified cyclase in R. dauricum.

No MeSH data available.