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Lipids

, Volume 35, Issue 3, pp 249–255 | Cite as

Cloning and characterization of the Dictyostelium discoideum cycloartenol synthase cDNA

  • Sharotka M. Godzina
  • Martha A. Lovato
  • Michelle M. Meyer
  • Kimberly A. Foster
  • William K. Wilson
  • Wei Gu
  • Eugenio L. de Hostos
  • Seiichi P. T. Matsuda
Article

Abstract

Cycloartenol synthase converts oxidosqualene to cycloartenol, the first carbocyclic intermediate en route to sterols in plants and many protists. Presented here is the first cycloartenol synthase gene identified from a protist, the cellular slime mold Dictyostelium discoideum. The cDNA encodes an 81-kDa predicted protein 50–52% identical to known higher plant cycloartenol synthases and 40–49% identical to known lanosterol synthases from fungi and mammals. The encoded protein expressed in transgenic Saccharomyces cerevisiae converted synthetic oxidosqualene to cycloartenol in vitro. This product was characterized by 1H and 13C nuclear magnetic resonance and gas chromatography-mass spectrometry. The predicted protein sequence diverges sufficiently from the known cycloartenol synthase sequences to dramatically reduce the number of residues that are candidates for the catalytic difference between cycloartenol and lanosterol formation.

Keywords

Lanosterol Dictyostelium Discoideum Euglena Gracilis Cycloartenol Triterpene Alcohol 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Abbreviations

A

alanine

C

cysteine

D

aspartate

E

glutamate

F

phenylalanine

G

glycine

H

histidine

I

isoleucine

K

lysine

L

leucine

M

methionine

N

asparagine

P

proline

Q

glutamine

R

arginine

S

serine

T

threonine

V

valine

W

tryptophan

Y

tyrosine

AaSHC

Alicyclobacillus acidocaldarius squalene-hopene cyclase

cDNA

complementary deoxyribonucleic acid

GC/MS

gas chromatography/mass spectrometry

HSQC

heteronuclear single quantum coherence

kbp

kilobase pair

NMR

nuclear magnetic resonance

PCR

polymerase chain reaction

PEG

polyethylene glycol

SC-UHET+gal

synthetic complete medium lacking uracil with 2% galactose, 13 mg/L heme, 20 mg/L ergosterol, 0.5% Tween 80

ScERG7

Saccharomyces cerevisiae lanosterol synthase

SC-UHET+glu

synthetic complete medium lacking uracil with 2% glucose, 13 mg/L heme, 20 mg/L ergosterol, 0.5% Tween 80

SHC

squalene-hopene cyclase

TLC

thin-layer chromatography

YPDHET

1% yeast extract, 2% peptone, 2% glucose, 13 mg/L heme, 20 mg/L ergosterol, and 0.5% Tween 80

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Copyright information

© AOCS Press 2000

Authors and Affiliations

  • Sharotka M. Godzina
    • 1
  • Martha A. Lovato
    • 2
  • Michelle M. Meyer
    • 1
    • 2
  • Kimberly A. Foster
    • 1
  • William K. Wilson
    • 1
  • Wei Gu
    • 1
  • Eugenio L. de Hostos
    • 1
  • Seiichi P. T. Matsuda
    • 1
    • 2
  1. 1.Department of Biochemistry and Cell BiologyRice UniversityHoustonTexas
  2. 2.Department of ChemistryRice UniversityHouston

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