* electrophilic addition results tertiary cation PPO * FPP 1' 2' 3' squalene synthase FPP allylic cation OPP HH* * loss of proton with formation of cyclopropane ring H * OPP * H presqualene PP loss of diphosphate results in primary cation * H W-M 1,3-alkyl shift * H * * H H 1,3-alkyl shift generates new bond cleavage produces cyclopropane ring and more alkene and favorable favorable tertiary cation allylic cation HH * * * * H H (NADPH) cation quenched by attack of anhydride Figure by MIT OCW. squalenesqualene O2 NADPH cyclizations O squalene oxide protonation of epoxide electrophilic addition electrophilic addition allows ring opening to gives tertiary cation + gives tertiary cation + tertiary cation six-membered ring six-membered ring H HOHOHO O H H squalene oxide H electrophilic addition gives tertiary cation + five-membered ring H H H H Figure by MIT OCW H H H HO H electrophilic addition H W-M rearrangement; HO HHO gives tertiary cation ring expansion despite protosteryl cation tertiary --> secondary cationH O HO H HH chair - boat - chair - boat squalene oxide HH sequence of W-M 1,2-hydride and HO HHO H 1,2-methyl shifts HH HH protosteryl cation sequence of W-M 1,2-hydride and HHH 1,2-methyl shifts HHH HO H HO H cyclopropane ring H HH protosteryl cation loss of H-9 creates formation and double bond loss of proton H from C-10 methyl H H HO H HO H H H HH H HO H cycloartenol HO H lanosterol Figure by MIT OCWAromatase-catalyzed conversion of androstenedione to estrone O OO O HO H CH3 HOH R HS HO OH O2 O2 -H2O H NADPH O NADPH OO O +H+ +H+ NADPH -H2O -HOH O2 +H+ O +H2O HCOOH + HO A Three possible mechanisms for the last step in the aromatase-catalyzed oxygenation of androstenedione. OO Fe+3 +3Fe O +3Fe O OOH O OOH +3Fe O H +3Fe OH 1 A O O O O +4Fe O O +4Fe OH O H O 2 A+ +4Fe OH C H Fe3+ + HCOOH HO HO O+4Fe O O +4Fe O CH O +4Fe O O +4Fe O O HH CH Fe3+ A+ + HCOOH 3 +4Fe OH HOOOO Figure by MIT OCW.PLANT HO HOOOH OH O H C-24 alkylation H loss of C methyl -4α H opening of cyclopropane HO H HO H HO H HO H loss of C-14α methyl and c y c l o a r t en o l 24-met h ylen ec y c lo ar t a n o l c y c lo eu c a len o l o b t u s ifo l i o l allylic isomerization (migration of Δ8 to Δ7) loss of C-4 further side-24-ethyl ste r o ls H H methyl H H chain alkylation H H loss of C-4 methyl H H HO H HO H HO H HO H epis ter o l av e n a s t e r o l c itrostad i e n ol gr a mi s t e r o l ( 2 4-et hy liden elo ph e n o l) ( 24-m e th y l en elo p h e n o l) 24-methyl sterols YEAST C-24 alkylation C-14 demethylation demethylations as C-4 H H side chain and HO HO HO HOH H HH ring B modifications lanosterol eburicol 4,4-dimethylfecosterol fecosterol (24-methylenedihydrolanosterol) Figure by MIT OCW. H H HO ergosterolcyclization is initiated by protonation of a double bond resulting in tertiary cation H+ squalene rings A and B are formed as 5-membered rings via Markovnikov addition; they then expand to 6-membered rings via W-M rearrangements H2O A B H2O OH H H H H tetrahymanol H H H H H H H H H hopan-22-ol OH HO Figure by MIT OCWGGPP PPO GGPP electrophilic addition allylic cation resulting in tertiary cation OPP H H loss of proton with formation of cyclopropane ring loss of diphosphate OPP results in primary H cation H H W-M 1,3-alkyl shift generates prephytoene PP new cyclopropane ring and more favorable tertiary cation HHH HH bond cleavage produces proton loss alkene and favorable generates alkene allylic cation H Z-phytoene Z sequence of desaturation reactions; in plants and fungi, the central double bond is also isomerized Z --> E E Figure by MIT OCW. lycopenec protonation of double bond H+ results in tertiary cation; then electrophilic addition O2 NADPH HOa ε-ring H bb H opening of pinacol-like rearrangement a γ-ring epoxide ring generates ketone H c O O2 O2 NADPH NADPH H+ HOHO HO β-ring O2 OH NADPH H allene generated from opening of epoxide O ring and loss of HO H+ HO OH proton OH O O O oxidative chain shortening central cleavage (a) excentric cleavage (b) excentric cleavage can generate one molecule of retinal central cleavage generates two molecules of retinal β-carotene (a) (b) Figure by MIT OCW. retinalAbietadiene Synthase bifunctional diterpene cyclase from fir cannot separate functional domains cloned (a.a. sequence is) Figure removed due to copyright reasons. don't know structure "homology modeling' ep.-aristolochene synthase enzyme as a model (sequiterpene synthase)Gibberrellin phytohormone bifunctional enzyme in fungi two enzymes in plant 2 individual enzymes OPPCPS KS FCPS/KSFCPS/KS Higher plants Figure by MIT OCW. Phaeosphaeria sp. L487 protonation ionization yeast 1 bifunctional enzyme Figure by MIT OCW. OPPIndex of figures removed due to copyright reasons Jennewein, S., and R. Croteau. Figure 2 in “Taxol: biosynthesis, molecular genetics, and biotechnological applications.” Applied Microbiol Biotech 57 (2001): 13-19. Jennewein, Stefan et al. “Random sequencing of an induced Taxus cell cDNA library for identification of clones involved in Taxol biosynthesis.” PNAS 101 (2004): 9149-9154. Brown, Geoffrey D. Scheme 8 in “The biosynthesis of steroids and triterpenoids.” Natural Product Reports 15 (1998): 653-696. Abe, Ikuro, Michel Rohmer, and Glenn D. Prestwich. Scheme VI in “Enzymatic cyclization of squalene and oxidosqualene to sterols and triterpenes.” Chemical Reviews 93 (1993): 2189 – 2206. Rilling, H.C., and Chayet, L.T. “The 19-reaction conversion of lanosterol to cholesterol.” Danielsson, H., and Sjovall, J., eds. Sterols and Bile Acids. New York, NY: Elsevier, 1985. ISBN: 0444806709. Hoshino, Tsutomu, and Tsutomu Sato. Figure 3 in “Squalene–hopene cyclase: catalytic mechanism and substrate recognition.” Chemical Communications (2002): 291 – 301. Peters, Reuben J. Scheme 1, Figure 1, and Table 1 in “Bifunctional Abietadiene Synthase: Mutual Structural Dependence of the Active Sites for Protonation-Initiated and Ionization-Initiated Cyclizations.” Biochemistry 42 (2003): 2700-2707. Williams, David C. et al. Figures 1, 3 and 6 in “Intramolecular proton transfer in the cyclization of geranylgeranyl diphosphate to the taxadiene precursor of taxol catalyzed by recombinant taxadiene synthase.” Chemistry & Biology 7 (2000): 969 -