PMID- 18324728
OWN - NLM
STAT- MEDLINE
DCOM- 20080731
LR  - 20080408
IS  - 0173-0835 (Print)
IS  - 0173-0835 (Linking)
VI  - 29
IP  - 7
DP  - 2008 Apr
TI  - The monitoring of nucleotide diphosphate kinase activity by blue native 
      polyacrylamide gel electrophoresis.
PG  - 1484-9
LID - 10.1002/elps.200700697 [doi]
AB  - Nucleoside diphosphate kinase (NDPK) has been shown to play a pivotal role in 
      modulating a plethora of cellular processes. In this study, we report on a blue 
      native (BN) PAGE technique which allows the facile assessment of NDPK activity 
      and expression. The in-gel detection of NDPK relies on the precipitation of 
      formazan at the site of immobilized enzyme activity. This is achieved by coupling 
      the formation of ATP, as a consequence of gamma-phosphate transfer from NTP to 
      ADP, to hexokinase (HK), glucose-6-phosphate dehydrogenase (G6PDH), oxidized 
      nicotinamide adenine dinucleotide phosphate (NADP), phenazine methosulfate (PMS), 
      and iodonitrotetrazolium chloride (INT). 2-D denaturing gel analysis confirmed 
      that the activity bands corresponded to NDPK as indicated by subunit composition. 
      Furthermore, the sensitivity and specificity of this readily accessible procedure 
      was assessed by monitoring the in-gel activity of NDPK using different 
      concentrations of GTP and CTP as well as deoxynucleoside triphosphates. This 
      electrophoretic technique allows the quick and easy detection of NDPK, a 
      housekeeping enzyme crucial to cell survival.
FAU - Mailloux, Ryan J
AU  - Mailloux RJ
AD  - Department of Chemistry and Biochemistry, Laurentian University, Sudbury, 
      Ontario, Canada.
FAU - Darwich, Rami
AU  - Darwich R
FAU - Lemire, Joseph
AU  - Lemire J
FAU - Appanna, Vasu
AU  - Appanna V
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Germany
TA  - Electrophoresis
JT  - Electrophoresis
JID - 8204476
RN  - EC 2.7.- (Phosphotransferases)
SB  - IM
MH  - Electrophoresis, Polyacrylamide Gel/*methods
MH  - Phosphotransferases/*metabolism
MH  - Pseudomonas fluorescens/enzymology
EDAT- 2008/03/08 09:00
MHDA- 2008/08/01 09:00
CRDT- 2008/03/08 09:00
PHST- 2008/03/08 09:00 [pubmed]
PHST- 2008/08/01 09:00 [medline]
PHST- 2008/03/08 09:00 [entrez]
AID - 10.1002/elps.200700697 [doi]
PST - ppublish
SO  - Electrophoresis. 2008 Apr;29(7):1484-9. doi: 10.1002/elps.200700697.