LytB protein catalyzes the terminal step of the 2-C-methyl-D-erythritol-4-phosphate pathway of isoprenoid biosynthesis.
Recombinant LytB protein from the thermophilic eubacterium Aquifex aeolicus produced in Escherichia coli was purified to apparent homogeneity. The purified LytB protein catalyzed the reduction of (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) in a defined in vitro system. The reaction products were identified as isopentenyl diphosphate and dimethylallyl diphosphate. A spectrophotometric assay was established to determine the steady-state kinetic parameters of LytB protein. The maximal specific activity of 6.6+/-0.3 micromol x min(-1) x mg(-1) protein was determined at pH 7.5 and 60 degrees C. The k(cat) value of the LytB protein was 3.7+/-0.2 s(-1) and the K(m) value for HMBPP was 590+/-60 microM.
Altincicek
Boran
B
Jomaa Pharmaka GmbH, Frankfurter Strasse 50, 35392, Giessen, Germany. boran.altincicek@jomaa.de
Duin
Evert C
EC
Reichenberg
Armin
A
Hedderich
Reiner
R
Kollas
Ann-Kristin
AK
Hintz
Martin
M
Wagner
Stefanie
S
Wiesner
Jochen
J
Beck
Ewald
E
Jomaa
Hassan
H
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004896:Erythritol / Q000031:analogs & derivatives* / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D005051:Eubacterium / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010755:Organophosphates / Q000494:pharmacology; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D013403:Sugar Phosphates / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors
C114232:2-C-methylerythritol 4-phosphate; C467061:4-hydroxy-3-methyl-2-butenyl diphosphate; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D010755:Organophosphates; D011994:Recombinant Proteins; D013403:Sugar Phosphates; D010088:Oxidoreductases; C109866:ispH protein, E coli; D004896:Erythritol