Directed evolution experiments reveal mutations at cycloartenol synthase residue His477 that dramatically alter catalysis.
[reaction: see text] Cycloartenol synthase cyclizes and rearranges oxidosqualene to the protosteryl cation and then specifically deprotonates from C-19. To identify mutants that deprotonate differently, randomly generated mutant cycloartenol synthases were selected in a yeast lanosterol synthase mutant. A novel His477Asn mutant was uncovered that produces 88% lanosterol and 12% parkeol. The His477Gln mutant produces 73% parkeol, 22% lanosterol, and 5% Delta(7)-lanosterol. These are the most accurate lanosterol synthase and parkeol synthase that have been generated by mutagenesis.
Segura
Michael J R
MJ
Department of Chemistry, Rice University, Houston, Texas 77005, USA.
Lodeiro
Silvia
S
Meyer
Michelle M
MM
Patel
Akash J
AJ
Matsuda
Seiichi P T
SP
D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000235:genetics; D002384:Catalysis; D017124:Conserved Sequence; D019020:Directed Molecular Evolution; D018698:Glutamic Acid / Q000235:genetics; D006639:Histidine / Q000235:genetics* / Q000378:metabolism*; D019751:Intramolecular Transferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007810:Lanosterol / Q000031:analogs & derivatives* / Q000378:metabolism; D015394:Molecular Structure; D009154:Mutation / Q000235:genetics*; D013329:Structure-Activity Relationship; D015003:Yeasts / Q000201:enzymology / Q000235:genetics
D007810:Lanosterol; C093882:parkeol; D001224:Aspartic Acid; D018698:Glutamic Acid; D006639:Histidine; D019751:Intramolecular Transferases; C021924:lanosterol synthase; C021923:cycloartenol synthase