PMID- 11128870
OWN - NLM
STAT- MEDLINE
DCOM- 20010208
LR  - 20190727
IS  - 0040-8727 (Print)
IS  - 0040-8727 (Linking)
VI  - 192
IP  - 1
DP  - 2000 Sep
TI  - Simple and rapid determination of Gtpase activity by capillary electrophoresis 
      without radioisotope.
PG  - 67-79
AB  - In order to determine guanosine-5'-triphosphatase (GTPase) activity, we developed 
      a simple, rapid and reliable method that utilizes capillary electrophoresis 
      without radioisotope. Tubulin-GTPase was used for simple measurement of GTPase 
      activity utilizing capillary electrophoresis. Tubulin, a component of 
      microtubules, was incubated with guanosine-5'-triphosphate (GTP) in 100 mM 
      2-(N-morpholino) ethanesulfonic acid (MES) buffer (pH 6.5). 
      Guanosine-5'-diphosphate (GDP) was determined as the hydrolyzed product of GTP. 
      Guanosine-5'-monophosphate, GDP and GTP in the filtrate of the mixture were 
      clearly separated using 10 mM MES buffer (pH 6.5) (migration time, 3.8, 5.5 and 
      7.2 minutes, respectively) with a fused-silica capillary column. The 
      quantification of GDP was based on the peak area, which increased linearly with 
      the concentration of GDP from 1 to 50 microM (r2=0.995). The peak area and 
      migration time had good reproducibility; the intra-assay coefficient of variation 
      (n=6) was 1.3% for peak area and 0.6% for migration time. As an application of 
      this method, we examined the effect of dimethylarsinic acid, an effective 
      antimitotic agent, on tubulin-GTPase. Dimethylarsinic acid inhibited 
      tubulin-GTPase activity in a dose-dependent manner. The inhibition was not 
      complete and the maximum decrease of the activity was about 50% at 200 microM 
      dimethylarsinic acid. Thus, since this method is clean, simple and rapid, its 
      application to the study of various GTPase proteins is expected to be useful.
FAU - Kawata, H
AU  - Kawata H
AD  - Department of Preventive Medicine and Environmental Health, Osaka City University 
      Medical School, Japan. hirozou@med.osaka-cu.ac.jp
FAU - Kuroda, K
AU  - Kuroda K
FAU - Endo, Y
AU  - Endo Y
FAU - Inoue, Y
AU  - Inoue Y
FAU - Endo, G
AU  - Endo G
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Japan
TA  - Tohoku J Exp Med
JT  - The Tohoku journal of experimental medicine
JID - 0417355
RN  - 0 (Buffers)
RN  - 0 (Tubulin)
RN  - 146-91-8 (Guanosine Diphosphate)
RN  - AJ2HL7EU8K (Cacodylic Acid)
RN  - EC 3.6.1.- (GTP Phosphohydrolases)
SB  - IM
MH  - Buffers
MH  - Cacodylic Acid/pharmacology
MH  - Electrophoresis, Capillary
MH  - GTP Phosphohydrolases/*metabolism
MH  - Guanosine Diphosphate/metabolism
MH  - Tubulin/metabolism
EDAT- 2000/12/29 11:00
MHDA- 2001/03/03 10:01
CRDT- 2000/12/29 11:00
PHST- 2000/12/29 11:00 [pubmed]
PHST- 2001/03/03 10:01 [medline]
PHST- 2000/12/29 11:00 [entrez]
AID - 10.1620/tjem.192.67 [doi]
PST - ppublish
SO  - Tohoku J Exp Med. 2000 Sep;192(1):67-79. doi: 10.1620/tjem.192.67.