PMID- 23383514
OWN - NLM
STAT- MEDLINE
DCOM- 20130319
LR  - 20130206
IS  - 0001-6209 (Print)
IS  - 0001-6209 (Linking)
VI  - 52
IP  - 11
DP  - 2012 Nov 4
TI  - [Cloning and characterization of a new antibacterial target, 
      3,4-dihydroxy-2-butanone-4-phosphate synthase].
PG  - 1415-20
AB  - OBJECTIVE: Riboflavin, called generally vitamin B12, is the precursor of cofactor 
      flavin adenine dinucleotide (FAD) and flavin mononucleotide, which plays crucial 
      roles in the biosynthesis of organisms. Bacteria need to synthesize riboflavin to 
      maintain their survival and proliferation if they cannot obtain flavin from the 
      surroundings. 3,4-Dihydroxy-2-butanone4-phosphate synthase (DHBPs) is one of the 
      key enzymes in biosynthesis of riboflavin. In the presence of Mg2+, DHBPs can 
      degrade ribulose-5-phosphate (Ru5P) into formate and 
      3,4-dihydroxy-2-butanone-4-phosphate (DHBP). Potentially, these enzymes related 
      to biosynthesis and metabolism of riboflavin, including DHBPs, may serve as the 
      target for new antibacterial drug. In order to determine the three-dimensional 
      structure and screen small molecule of inhibitor of DHBPs, we expressed and 
      purified DHBPs from Streptococcus pneumonia (S. pn), and characterized the 
      activity of this enzyme. METHOD: DHBPs gene was amplified by PCR, and 
      over-expression plasmid pW28-DHBPs was constructed. pW28-DHBPs was transformed 
      into Escherichia coli BL21 (DE3) to express DHBPs; the recombinant protein was 
      purified by nickel column and ion-exchange column. The enzymatic activity was 
      tested using spectroscopy. RESULTS: The plasmid of pW28-DHBPs was verified by 
      restrictive enzyme digestion and sequencing. Soluble DHBPs was expressed in E. 
      coli BL21, and purified with 95% purity. The result of size exclusion 
      chromatography indicates that DHBPs was dimer in solution. Additionally, the 
      recombinant protein has activity to hydrolyze Ru5P into formate and DHBP in the 
      conditions of pH 7.5 and 25 degrees C and in the presence of Mg2+. CONCLUSIONS: 
      DHBPs could be highly expressed in soluble form in E. coli BL21 strain, and the 
      recombinant protein has activity to hydrolyze Ru5P.
FAU - Jin, Li
AU  - Jin L
AD  - Key Laboratory of Molecular Biology of Infectious Diseases, Ministry of 
      Education, Faculty of Laboratory Medicine, Chongqing Medical University, 
      Chongqing 400016, China. lijin2011@126.com
FAU - Zhou, Hua
AU  - Zhou H
FAU - Zhao, Shasha
AU  - Zhao S
FAU - Yang, Wei
AU  - Yang W
FAU - Niu, Siqiang
AU  - Niu S
FAU - Wang, Deqiang
AU  - Wang D
LA  - chi
PT  - English Abstract
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - China
TA  - Wei Sheng Wu Xue Bao
JT  - Wei sheng wu xue bao = Acta microbiologica Sinica
JID - 21610860R
RN  - 0 (Anti-Bacterial Agents)
RN  - 0 (Bacterial Proteins)
RN  - 0 (Enzyme Inhibitors)
RN  - 0 (Ribulosephosphates)
RN  - 4151-19-3 (ribulose 5-phosphate)
RN  - EC 5.4.- (Intramolecular Transferases)
RN  - EC 5.4.- (L-3,4-dihydroxy-2-butanone-4-phosphate synthase)
SB  - IM
MH  - Anti-Bacterial Agents/*pharmacology
MH  - Bacterial Proteins/antagonists & inhibitors/*chemistry/*genetics/metabolism
MH  - *Cloning, Molecular
MH  - Drug Resistance, Bacterial
MH  - Enzyme Inhibitors/pharmacology
MH  - Intramolecular Transferases/antagonists & 
      inhibitors/*chemistry/*genetics/metabolism
MH  - Ribulosephosphates/metabolism
MH  - Streptococcus pneumoniae/drug effects/*enzymology/genetics/metabolism
EDAT- 2013/02/07 06:00
MHDA- 2013/03/21 06:00
CRDT- 2013/02/07 06:00
PHST- 2013/02/07 06:00 [entrez]
PHST- 2013/02/07 06:00 [pubmed]
PHST- 2013/03/21 06:00 [medline]
PST - ppublish
SO  - Wei Sheng Wu Xue Bao. 2012 Nov 4;52(11):1415-20.